Open access peer-reviewed chapter

Development of an Oral Vaccine for the Control of Cysticercosis

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Marisela Hernández, Anabel Ortiz Caltempa, Jacquelynne Cervantes, Nelly Villalobos, Cynthia Guzmán, Gladis Fragoso, Edda Sciutto and María Luisa Villareal

Submitted: February 2nd, 2021 Reviewed: March 15th, 2021 Published: April 29th, 2021

DOI: 10.5772/intechopen.97227

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Parasitic diseases fecally transmitted, such taeniasis/cysticercosis Taenia solium binomial, represent a health problem whose incidence continues due to the prevalence of inadequate sanitary conditions, particularly in developing countries. When the larval stage of the parasite is established in the central nervous system causes neurocysticercosis a disease than can severely affect human health. It can also affect pigs causing cysticercosis causing economic losses. Since pigs are obligatory intermediate hosts, they have been considered as the targets for vaccination to interrupt the transmission of the parasitosis and eventually reduce the disease. Progress has been made in the development of vaccines for the prevention of porcine cysticercosis. In our research group, three peptides have been identified that, expressed synthetically (S3Pvac) or recombinantly (S3Pvac-phage), reduced the amount of cysticerci by 98.7% and 87%, respectively, in pigs exposed to natural conditions of infection. Considering that cysticercosis is orally acquired, it seems feasible to develop an edible vaccine, which could be administered by the pig farmers, simplifying the logistical difficulties of its application, reducing costs, and facilitating the implementation of vaccination programs. This chapter describes the most important advances towards the development of an oral vaccine against porcine cysticercosis.


  • cysticercosis
  • T. solium
  • oral vaccine
  • transgenic plant
  • Carica papaya

1. Introduction

Taenia solium taeniasis/cysticercosis is a parasitic zoonosis that significantly affect economic and public health. Neurocysticercosis (NCC) is a most severe form of the disease caused by the establishment of the larval stage (cysticerci) of Taenia solium in the central nervous system (CNS). In 2010, the World Health Organization declared it one of the leading neglected diseases and aims to develop strategies for its eradication and prevention [1].

Between control measures it has been explored the improvement of health education, sanitary conditions, standards of meat inspection and the rearing of pigs. It has also been explored the impact of massive or individual treatment of taeniasis and the treatment and/or vaccination of pigs, all of them with promising results [2, 3, 4]. Vaccination of pigs would imply an unlikely immediate and potent effect upon the number of tapeworm-carriers in rural communities, interrupting the parasite’s life cycle and eventually reduce human neurocysticercosis. Developing an effective vaccine against T. solium pig cysticercosis is also being pursued by different research groups with promising results [5, 6].

In our group, an anti-cysticercosis vaccine named S3Pvac based on three peptides expressed was developed. The vaccine synthetically (S3Pvac) or recombinant (S3Pvac-phage) produced, reduced the number of cysticerci by 98.7% and 87% [7, 8] in randomized field trials, respectively. The recombinant vaccine was subsequently used in a control program applied in the State of Guerrero, confirming its usefulness. Indeed, S3Pvac-phage significantly reduce the prevalence of porcine cysticercosis from 7 to 0.5% and 3.6 to 0.3% estimated by tongue examination or ultrasound, respectively [3]. In the course of this control program, we were able to evaluate the difficulties involved in using an injectable vaccine. Pigs are produced free rurally reared, thus the application of an injectable vaccine requires their capture and subjection, a laborious procedure that increases the costs of vaccination and limits its massive application. Considering that cysticercosis is orally acquired, it seems feasible to develop an oral vaccine [9], which could be administered by the pig breeder, simplifying the logistical difficulties of its administration, reducing costs and facilitating the implementation of vaccination control programs.

For the design of an oral vaccine the use of plants is increasingly recognized as valuable platform. Plants offer the production of antigens at low costs, circumventing costly purification procedures. Plants also offer a natural way of antigen encapsulation preventing its degradation by the detrimental environment to which an oral intake vaccine is exposed such us antigen degradation by low pH, mucosal enzymes [10, 11] and the use of cell cultures will avoid non-desirable environmental effects due to the release of transgenic plants into the environment.

Moreover, plants also frequently include components with adjuvant properties like saponins that may increase the immunogenicity of the vaccine [12]. Considering this, the recombinant peptides KETc7, KETc1.6His, and KETc12.6His were expressed in transgenic clones of papaya embryogenic calli [13]. The three clones together constitute the oral S3Pvac-papaya vaccine candidate. Papaya was selected because the high efficiency of transformation and its own antiparasitic properties [14].

This third version of the vaccine has been shown to be immunogenic in mice and pigs and is being produced in suspended culture systems to massively produced this oral version of the vaccine that must be evaluated on the field against pig cysticercosis.


2. Parasite and oral immunity

Oral vaccination is an interest route to prevent infections caused by orally acquired pathogens overcoming the limitations of current injection-based vaccines in providing front-line protection against pathogen invasion and dissemination [15]. It offers a painless, safe and low-cost route that does not require trained personnel. Moreover, this route can elicit mucosal and systemic immunity. Vaccine antigen can be recognized and translocated by M cells, which act as sentinels and enter directly into the Peyer’s patches. Then antigens can be transported to the intestinal mesenteric lymph nodes, stimulating the host’s systemic and mucosal immune response resulting in the production of IgA and IgG antibodies with the ability to neutralize of invading pathogens before they are able to cause a widespread infection. Oral vaccination can also trigger an effective cellular immunity. However, the development of oral vaccines is a major challenge due to an inefficient transport to reach M cells and the possibility to induce local and systemic immune tolerance. Considering that plant-based vaccines usually expressed low content of antigen it seems feasible to avoid oral tolerance using the proper dose and vaccine schedule. It remains to be elucidated if plants-derived vaccines could overcome mucosal tolerance when administered to human beings.

Table 1 shows various plants that have been used to express antigens from different pathogens to be evaluated as edible vaccines. Tobacco has been used as an experimental model of transformation and expression. However, the use of other species such as tomatoes, lettuce, potatoes, corn, soybeans, alfalfa, Arabidopsis, papaya and carrots has been expanded [11, 28, 29, 30, 31, 32, 33]. In some of these plants, the expressed recombinant antigen has shown efficacy when evaluated in experimental models or directly in the naturally affected host. Recombinant antigens have been reported to induce an immune response with the production of IgG, IgM or IgA antibodies, regardless of the route of administration [31].

Rabies virusTomatoGPNDND[16]
PotatoVP60Sc, ImRabbit[17]
Rotavirus APotatoVP6IpMice[18]
Foot and mouth virusArabidopsisVP1IpMice[19]
Foot and mouth virusPotatoVP1IpMice[20]
Foot and mouth virusAlfalfaPeptide VP1
Gastroenteritis virusArabidopsisGP- SImMice[22]
Gastroenteritis virusPotatoGP- SOralMice[23]
Gastroenteritis virusTobaccoGP- SIpPig[24]
Gastroenteritis virusSeeds of cornGP- SOralPig*[25, 26]
S3Pvac-papayaEmbryogenic Transgenic clonesKETc1, KETc12 and KETc7 peptidesScMice[13]
S3Pvac-papayaEmbryogenic Transgenic clonesKETc1, KETc12 and KETc7 peptidesOralPig[27]

Table 1.

Expression of antigens aimed at veterinary vaccine development.

Protection against infection.

Sc: subcutaneous; Im: intramuscular; Ip: intraperitoneal, ND: Not determined; GP: Glycoprotein.


3. Transgenic plant platform

Many different advantages of expression of recombinant proteins in transgenic plants for vaccine production can be mentioned over other commonly expression systems, such as bacteria, yeasts and baculoviruses. Plants can be constitutively or tissue-specific expressed in single or multiple transgenes, antigens can be stable in seeds without the requirement of refrigeration, no purification nor cold chain for preservation.

Transgenic plants can also be used as bioreactors to produce high amounts of the recombinant protein of interest [34, 35]. They can also be produced as in vitro tissue culture, cell suspensions, hairy roots, moss protonema, microalgae and whole plants. There are many experimental plant-made veterinary vaccines produced in seeds, fruits, and leaves, that can be orally delivered as part of the animal feed, thus offering great convenience and economy in immunizing large populations of animals on farms [35]. The expression of antigens for the production of vaccines in transgenic plants is considered a safe and effective immunization system, which can avoid some of the difficulties associated with traditional vaccination methods, as well as a reduction in the costs of production, distribution and conservation.

One nice study of veterinary interest is the expression of the glycoprotein S of the porcine gastroenteritis virus in corn seeds for the production of an oral vaccine, which has also the ability to induce protection, through colostrum, in piglets [25, 26].

3.1 Carica papaya L.

Classification of Carica papaya L.

Family: Caricaceae.

Gender: Carica.

Species: C. papaya L.

Morphological type: Arboreal.

Climate: Equatorial tropical.

Carica papaya is a species of pantropical plant that grow in tropical regions of the Americas from Mexico to Argentina, Africa and Asia. The main importers are: United States, Japan, Hong Kong and the European Union. Carica papaya is known by different common names such as capaidso, naimi, nampucha, pucha, fruit bomb, milky, mamao, pawpaw. Papaya is an arborescent, semi-perennial plant that grows in areas with an average rainfall of 1800 mm per year and an average annual temperature of 20–22°C, a large number of varieties have been developed. Papaya fruiting occurs 10 to 12 months after transplantation, is maintained for ten years, and female, male or hermaphrodite [36] trees can be obtained. Papaya is a fruit known for its nutritional benefits and medicinal properties. Main papaya components and their reported properties are shown in Table 2.

Food fiberCaricainStrengthens ImmunitySpermatocide
FatGlycyl endopeptidaseAntibacterialEmmenagogue
ProteinsLesser amountsContraceptive
Retinol (vit. A)Class II chitinaseAnalgesic
β-caroteneClass III chitinaseAntibiotic
Thiamine (vit. B1)Serin protease inhibitorStimulation of pancreatic juices
Riboflavin (vit. B2)Glutamyl cyclotransferaseHypotensive
Niacin (vit. B3)Beta-1,3-glucanaseFebrifuge
Pantothenic acid (vit. B5)CystatinAnti-inflammatory
Folic acid (vit. B9)Acetogenins*Anti-helminthic
Vitamin CCarpasemine
Vitamin E
Vitamin K

Table 2.

Papaya components and medicinal properties reported.

Antiparasitic activity. Calcium, iron, magnesium, manganese, phosphorus, potassium, sodium and Zn are also included in papaya.

3.2 Carica papaya as a cestode vaccine

Papaya is an alternative system for the exploration of tropical tree genomes, containing a genome of 372 megabase (Mb), of diploid inheritance with 9 pairs of chromosomes and presents the smallest gene number, 24,746 genes [37]. Papaya exhibits some properties of possible advantages to be used as a platform to express T. solium vaccine antigens. Papaya components have antiparasitic properties per se [14]. Cells can be easily transformed by bioballistics and in vitro propagated and regenerated [38].

Among many papaya components, the papain contained in the latex has been widely evaluated in its ability to damage the cuticle of intestinal parasites by proteolytic digestion. Table 3 shows some reports on the characterization and evaluation of antiparasitic activity of papaya against Trichostrongylus colubrormis, Heligmosomoides polygyrus, Trichuris muris, Protospirura muricola [3944, 45, 46, 47, 48, 49] Rodentolepis microstoma [39], Hymenolepis diminuta and microstoma [40]. Anoplocephala perfoliate [41] Hymenolepis diminuta [42] Hymenolepis microstoma [43] without causing side effects to the host [50, 51].

In vitro
Rodentolepis microstomaPapain
Raw latex (Sigma)
25 mMDisrupt the surface of the cuticle[39]
Hymenolepis diminuta
Latex supernatant
Commercial papain
Not availableDamage to the strobile
Reduced motility and subsequent death of the parasite
Anoplocephala perfoliataCysteine-proteinases
Latex supernatant
300 mMReduced motility and induced death of the parasite[41]
Hymenolepis diminutaPapaya latex supernatant2.4 μmolAffected worm growth[42]
In vivo(mouse)
Hymenolepis microstomaLatex supernatant240 nm for 6 daysMinimal and temporary efficacy[43]

Table 3.

Antiparasitic activity of papaya against some cestodes.

3.3 S3Pvac-papaya anticysticercosis vaccine

For the development of the S3Pvac-papaya cysticercosis vaccine, three genetic constructions were used for the expression of recombinant peptides, KETc1.6His, KETc12.6His and KETc7 [13]. Figure 1 summarize the methodology employed for the production of S3Pvac-papaya vaccine.

Figure 1.

(A) Production of transgenic embryogenic papaya clones by a bioballistic method (B) Embryogenic papaya callus: a) Induction of embryogenic callus; b) Embryos in globular stage for transformation; c) Selection of transformed clones in selective medium.

3.4 Protective properties of S3Pvac papaya against cysticercosis

The S3Pvac vaccine expressed in embryogenic papaya clones has demonstrated high protective capacity against experimental murine and T. pisiformis cysticercosis orally administered. Table 4 shows the protective effect induced by oral immunization in mice at a dose range of 0.1 to 1 μg of soluble extract, whilst a higher dose lowered the percent of protection. In addition, different vaccine formulations also reduced the expected parasite load. On the other hand, the oral vaccine significantly reduced the number of infected rabbits and the percentage of cysticercus-free animals (83%), 21 days after the infection. The S3Pvac-papaya vaccine has not yet been evaluated in pigs, however, its immunogenic response in mice and pigs [27, 52], and its protective capacity in different models exhibit its potential to exert a protective response against naturally acquired porcine cysticercosis.

Experimental modelImmunized with:Mean ± SD (% Protection)Immune responseReference
Saline27.2 ± 14.2
S3Pvac papaya saline (μg/dose)
T. crassiceps
0.112.3 ± 3.4* (55)CD4, CD8 proliferation
110.8 ± 2.5* (60)CD4, CD8 proliferation
109.2 ± 1.0*
Specific IgG Abs; CD4 and CD8 proliferation
10041.4 ± 62
Saline18.6 ± 17.3* (86.8)
Corn starch§29.8 ± 31.4 (41.4)[27]
Soy oil10.3 ± 2.2* (75)
Canola oil11.5 ± 8.6* (84)
Rabbit experimental T. pisiformisSaline4.33 ± 4.01[53]
S3Pvac papaya£0.25 ± 0.62*

Table 4.

Protective capacity induced by oral S3Pvac-papaya vaccine against experimental.

Mean ± standard deviation of the number of cysticerci recovered in each group.

Mice were fed with S3Pvac-papaya soluble extract (1 μg of total protein) into different vehicles.

Rabbits received a suspension of 20 mg of each embryogenic transgenic papaya clone expressing KETc1, KETc12 and KETc7 in a gelatin capsule.

Protection statistically significant (P < 0.05).

We previously reported non-specific protection that was induced by the wild type soluble extract [52, 54] has been attributed to the antiparasitic properties described to papaya itself mentioned above.

3.5 Biotechnological approach for the production of papaya anti-cysticercosis vaccine

Plant biotechnology is a rapidly evolving area with major impact in the production of molecules with high pharmaceutical value. In vitro culture techniques offer central advantages in the manufacture of desired chemicals for human health. The benefits include a systematic supply of compounds under optimized controlled conditions, independence of weather, soil, disease, and socio-political problems; discovery of new compounds, bio-transformation systems and better adaptation to market changes. In an inclusive context, in vitro systems will give a better understanding of plant biochemistry and physiology, as well as some basic aspects of plant differentiation.

Plant biotechnology involves relevant procedures in the manufacture of oral vaccines enabling the production of higher amounts of active biomasses from transgenic plants, by means of massive propagation of cells, tissues and organs [1255]. Among others, these procedures include the growth of callus (aggregates of undifferentiated cells growing in solid media), suspension cultures (individualized undifferentiated cells growing in liquid media); as well as embryo cultures that could be grown in solid or in liquid nutrient media.

The three callus lines expressing KETc1, KETc7 and KETc12 peptides were generated, and further efforts to optimize the massive growth of the corresponding callus and suspension cultures, were conducted. These in vitro systems constitute adequate platforms for the massive production of papaya anti-cysticercosis vaccine in the near future.

3.5.1 Callus cultures

In the establishment and optimization of callus cell lines, Carica papaya L. (KETc7) embryogenic calli were used to obtain friable undifferentiated cells. Calli were placed in solid culture medium with 30 g/L sucrose, 3 g/Lpolivinilpilorridone and 1.5 g/Lof phytagel. The nutrient media MS [56] and B5 [57] were evaluated; and the presence of the phytoregulators 2,4-dichlorophenoxyacetic acid (2,4-D) at 1.0, 2.0 and 3.0 mg/Lcombined with kinetin (KN) at 1.0, 2.0 and 3.0 mg/L, was also tested. The cultures were maintained at 25°C and subjected to constant light as well as dark conditions. The best results were obtained for callus growing in B5 medium with 2 mgL−1 of 2,4-D combined with 2 mgL−1 KN in dark conditions (Figure 2). In these conditions after two subcultures non-phenolized calluses were developed, and after several subcultures the friable callus KETc7 cell line, was established.

Figure 2.

Optimization of C. papaya KETc7 callus cell line under different growth conditions. (a) Photoperiod, (b) constant light, (c) darkness.

3.5.2 Cell suspension cultures

Ten grams fresh weight (FW) of the friable callus line KETc7 were inoculated in 250 ml Erlenmeyer baffled flasks containing 100 ml nutrient medium without phytagel, and placed for 30 days on a rotary shaker at 100 rpm, 25°C and dark conditions. To disaggregate cell clusters and increase oxygen transfer, baffled flasks were used (Figure 3).

Figure 3.

Establishment of C. papaya KETc7 cell suspension line. (a) Culture in baffle flasks at 15 days (b) culture in Erlenmeyer flasks at 15 days (c) culture in Erlenmeyer flasks at 30 days.

The cultures were sub-cultured in fresh medium every 15 days, and the best results were observed when using B5 nutrient medium, cultivated in darkness, and producing uniform suspended cultures without phenolization (Figure 4).

Figure 4.

Growth of C. papaya KETc7 cell suspension line at 15 days in a rotary shaker at 100 rpm, 25°C, in the dark.

Cell viability was maintained at 95% until 45 days in culture, as confirmed by the fluorescein diacetate (FDA) method (Figure 5) [58].

Figure 5.

Cell viability of C. papaya KETc7 cell suspension line by the fluorescein diacetate (FDA) method at day a) 0, b) 7, and c) 15, in an Epifluorescence Microscope Nikon Eclipse E400 (40X).

Once the friable uniform cell suspension cell line was established, it became possible to evaluate the growth kinetics of the culture during 45 days by collecting samples every 3 days, and determining the following growth parameters: fresh weight, dry weight, cell viability, pH and carbohydrate consumption [59].

The results showed that the KETc7 suspension cell line grew very well, reaching a doubling time of 6.9 days and a specific growth rate (μ) of 0.10 d−1. The maximum biomass value was 14.36 gPS L−1 obtained at 15 days in culture.

3.5.3 Embryo suspension cultures

The KETc7 embryos callus cell line generated in solid B5 medium without phytoregulators was used to establish embryo suspension cultures. An inoculum of 10% was added into 250 ml Erlenmeyer flasks containing 100 ml of liquid B5 medium (Figure 6). The flasks were kept on an orbital shaker at a stirring speed of 115 rpm, under constant light conditions (24 μmol.m−2. S−1) and 25°C. After 15 days, the biomass was sub cultivated in the same conditions described above, and the culture was propagated.

Figure 6.

C. papaya KETc7 embryo suspension line grown in B5 medium at 100 rpm, 25°C, and constant light (24 μmol.m−2. S−1).

3.5.4 Cell suspension cultures in bioreactors

To scale-up the C. papaya suspension cultures 2 L airlift bioreactors were employed, with the following geometric design: height (52 cm), diameter (7 cm), draft tube height (27 cm), diameter of inner draft tube (2.7 cm), and bottom clearance (2.0 cm) [60]. The air was sprayed at the bottom of the draft, generating an internal loop in which the upcomer is in the draft, and the downcomer in the ring. The bioreactor was sterilized and then filled with autoclaved B5 medium (1.8 L) supplemented with 30 g/L sucrose, 2,4-D and KN (2 mg/L each). Fifteen-days-old C. papaya KETc7 cell suspension line was used to obtain an inoculum of 10% (v/v) FW. The culture in bioreactor was incubated at 25 ± 2°C under continuous light (white light flux density of 50 μmol/m2/s) for 30 days. The bioreactor was operated in a batch mode at 0.1 vvm for 15 days and subsequently at 0.8 vvm, until the end of the culture period. Under these conditions, an adequate mixing of the cell suspension was obtained. Antifoam (Dow Corning FG-10) was applied as required, by injection of 0.5 mL (0.1% v/v). The culture was sampled every three days and the concentration of biomass, pH and sugars, was determined. Results show that the K ETc7 cell suspension culture was able to grow uniformly and that the exponential growth phase was reached from days 6 to 12, followed by a stationary phase. The maximum biomass was of 18.6 ± 0.7 g/L DW (Figure 7).

Figure 7.

C. papaya KETc7 cell suspension line growing in airlift bioreactor: (a) day 0, (b) day 15, (c) day 30.

3.5.5 Embryo suspension cultures in bioreactors

Growth of C. papaya KETc7 embryo suspension line was scaled-up in the 2 L airlift bioreactors described before. Two weeks old embryo suspension line was used to obtain an inoculum of 10% (v/v) FW. The culture in bioreactor was incubated at 25 ± 2°C under continuous light (white light flux density of 50 μmol/m2/s) for 30 days. The bioreactor was operated in a batch mode at 0.1 vvm for 15 days and subsequently at 0.8 vvm until the end of the culture period. Embryo culture of the line KETc7in bioreactor batch type process, showed uniform growth. A maximum biomass of 30 g/L DW was obtained, which represents 4 times more respect to the initial inoculum and the number of generated embryos was of 279 (Figure 8).

Figure 8.

C. papaya KETc7 embryo suspension line growing in airlift bioreactor (a) embryo culture in airlift bioreactor at 30 days, (b) harvested embryos after 30 days in culture, (c) embryos observed in stereoscopic microscope (10×).


4. Conclusions

This review addresses oral vaccination as a feasible approach to prevent parasitic diseases. Since most anti-parasitic vaccines currently available are parenterally administered, their use involves high production and logistic costs and become inaccessible for underdeveloped countries. To cope with this issue, the use of papaya transgenic clones is herein proposed to develop an anti-cysticercosis oral vaccine and to assay its effectiveness against other parasitic infections of veterinary and/or public health interest. The use of biotechnological tools by escalation of suspension cultures would allow us to produce a vaccine in sufficient, controlled amounts for its direct application, reducing the use of antibiotics, and therefore the risk of bacterial resistance.



Thanks to Georgina Díaz, B.Sc., and Gonzalo Acero, B.Sc., for their technical support. Special thanks to Juan Francisco Rodríguez, B.Sc., for his help in preparing the original manuscript. This work was supported by the Programa de Investigación para el Desarrollo y la Optimización de Vacunas, Inmunomoduladores y Métodos Diagnósticos, Institute of Biomedical Research, UNAM.


Conflict of interest

The authors declare no conflict of interest.


  1. 1. WHO. The Control of Neglected Zoonotic Diseases-Report of the third conference organized with ICONZ, DFID-RIU, Gates Foundation, SOS, EU, TDR and FAO with the participation of ILRI and OIE WHO headquarters, Geneva, Switzerland 23-24 november 2010. 2010
  2. 2. Gabriël S, Dorny P, Mwape KE, Trevisan C, Braae UC, Magnussen P et al. Control of Taenia solium taeniasis/cysticercosis: The best way forward for sub-Saharan Africa? Acta Trop 2017; 165: 252-260
  3. 3. De Aluja AS, Suárez-Marín R, Sciutto-Conde E, Morales-Soto J, Martínez-Maya JJ, Villalobos N. Evaluación del impacto de un programa de control de la teniasis-cisticercosis (Taenia solium) [Evaluation of the impact of a control program against taeniasis-cysticercosis (Taenia solium)]. Salud Publica Mex 2014; 56: 259
  4. 4. Garcia HH, Gonzalez AE, Tsang VCW, O’Neal SE, Llanos-Zavalaga F, Gonzalvez G et al. Elimination of Taenia solium Transmission in Northern Peru. N Engl J Med 2016; 374: 2335-2344
  5. 5. Gauci C, Jayashi C, Lightowlers MW. Vaccine development against the taenia solium parasite: The role of recombinant protein expression in Escherichia coli. Bioengineered 2013; 4. doi:10.4161/bioe.23003
  6. 6. Sciutto E, Fragoso G, Hernández M, Rosas G, Martínez JJ, Fleury A et al. Development of the S3Pvac vaccine against murine taenia crassiceps cysticercosis: A historical review. J Parasitol 2013; 99: 693-702
  7. 7. Huerta M, De Aluja AS, Fragoso G, Toledo A, Villalobos N, Hernández M et al. Synthetic peptide vaccine against Taenia solium pig cysticercosis: Successful vaccination in a controlled field trial in rural Mexico. Vaccine 2001; 20: 262-266
  8. 8. Morales J, Martínez JJ, Manoutcharian K, Hernández M, Fleury A, Gevorkian G et al. Inexpensive anti-cysticercosis vaccine: S3Pvac expressed in heat inactivated M13 filamentous phage proves effective against naturally acquired Taenia solium porcine cysticercosis. Vaccine 2008; 26: 2899-2905
  9. 9. Cripps AW, Kyd JM, Foxwell AR. Vaccines and mucosal immunisation. In: Vaccine. Vaccine, 2001, pp. 2513-2515
  10. 10. Mason G, Provero P, Vaira AM, Accotto GP. Estimating the number of integrations in transformed plants by quantitative real-time PCR. BMC Biotechnol 2002; 2. doi:10.1186/1472-6750-2-20
  11. 11. Walmsley AM, Arntzen CJ. Plants for delivery of edible vaccines. Curr Opin Biotechnol 2000; 11: 126-129
  12. 12. Rosales-Mendoza S, Salazar-González JA. Immunological aspects of using plant cells as delivery vehicles for oral vaccines. Expert Rev Vaccines 2014; 13: 737-749
  13. 13. Hernández M, Cabrera-Ponce JL, Fragoso G, López-Casillas F, Guevara-García A, Rosas G et al. A new highly effective anticysticercosis vaccine expressed in transgenic papaya. Vaccine 2007; 25: 4252-4260
  14. 14. Stepek G, Behnke JM, Buttle DJ, Duce IR. Natural plant cysteine proteinases as anthelmintics? Trends Parasitol 2004; 20: 322-327
  15. 15. Miquel-Clopés A, Bentley EG, Stewart JP, Carding SR. Mucosal vaccines and technology. Clin Exp Immunol 2019; 196: 205-214
  16. 16. McGarvey PB, Hammond J, Dienelt MM, Hooper DC, Fang Fu Z, Dietzschold B et al. Expression of the rabies virus glycoprotein in transgenic tomatoes. Bio/Technology 1995; 13: 1484-1487
  17. 17. Castañón S, Marín MS, Martín-Alonso JM, Boga JA, Casais R, Humara JM et al. Immunization with Potato Plants Expressing VP60 Protein Protects against Rabbit Hemorrhagic Disease Virus. J Virol 1999; 73: 4452-4455
  18. 18. Matsumura T, Itchoda N, Tsunemitsu H. Production of immunogenic VP6 protein of bovine group A rotavirus in transgenic potato plants: Brief report. Arch Virol 2002; 147: 1263-1270
  19. 19. Carrillo C, Wigdorovitz A, Oliveros JC, Zamorano PI, Sadir AM, Gómez N et al. Protective Immune Response to Foot-and-Mouth Disease Virus with VP1 Expressed in Transgenic Plants. J Virol 1998; 72: 1688-1690
  20. 20. Carrillo C, Wigdorovitz A, Trono K, Dus Santos MJ, Castañón S, Sadir AM et al. Induction of a virus-specific antibody response to foot and mouth disease virus using the structural protein VP1 expressed in transgenic potato plants. Viral Immunol 2001; 14: 49-57
  21. 21. Dus Santos MJ, Wigdorovitz A, Trono K, Ríos RD, Franzone PM, Gil F et al. A novel methodology to develop a foot and mouth disease virus (FMDV) peptide-based vaccine in transgenic plants. Vaccine 2002; 20: 1141-1147
  22. 22. Gómez N, Carrillo C, Salinas J, Parra F, Borca M V., Escribano JM. Expression of immunogenic glycoprotein S polypeptides from transmissible gastroenteritis coronavirus in transgenic plants. Virology 1998; 249: 352-358
  23. 23. Gómez N, Wigdorovitz A, Castañón S, Gil F, Ordá R, Borca M V. et al. Oral immunogenicity of the plant derived spike protein from swine-transmissible gastroenteritis coronavirus. Arch Virol 2000; 145: 1725-1732
  24. 24. Tuboly T, Yu W, Bailey A, Degrandis S, Du S, Erickson L et al. Immunogenicity of porcine transmissible gastroenteritis virus spike protein expressed in plants. Vaccine 2000; 18: 2023-2028
  25. 25. Streatfield SJ, Jilka JM, Hood EE, Turner DD, Bailey MR, Mayor JM et al. Plant-based vaccines: Unique advantages. Vaccine 2001; 19: 2742-2748
  26. 26. Lamphear BJ, Streatfield SJ, Jilka JM, Brooks CA, Barker DK, Turner DD et al. Delivery of subunit vaccines in maize seed. J Control Release 2002; 85: 169-180
  27. 27. Fragoso G, Hernández M, Cervantes-Torres J, Ramírez-Aquino R, Chapula H, Villalobos N et al. Transgenic papaya: a useful platform for oral vaccines. Planta 2017; 245: 1037-1048
  28. 28. Floss DM, Falkenburg D, Conrad U. Production of vaccines and therapeutic antibodies for veterinary applications in transgenic plants: An overview. Transgenic Res 2007; 16: 315-332
  29. 29. Rosales-Mendoza S, Soria-Guerra RE, López-Revilla R, Moreno-Fierros L, Alpuche-Solís ÁG. Ingestion of transgenic carrots expressing the Escherichia coli heat-labile enterotoxin B subunit protects mice against cholera toxin challenge. Plant Cell Rep 2008; 27: 79-84
  30. 30. Sala F, Rigano MM, Barbante A, Basso B, Walmsley AM, Castiglione S. Vaccine antigen production in transgenic plants: Strategies, gene constructs and perspectives. In: Vaccine. Vaccine, 2003, pp. 803-808
  31. 31. Streatfield SJ, Howard JA. Plant-based vaccines. Int J Parasitol 2003; 33: 479-493
  32. 32. Streatfield SJ. Mucosal immunization using recombinant plant-based oral vaccines. Methods 2006; 38: 150-157
  33. 33. Tacket CO. Plant-derived vaccines against diarrheal diseases. Vaccine 2005; 23:1866-1869
  34. 34. Xu J, Towler M, Weathers PJ. Platforms for Plant-Based Protein Production. 2018 doi:10.1007/978-3-319-54600-1_14
  35. 35. Massa S, Presenti O, Benvenuto E. Engineering Plants for the Future: Farming with Value-Added Harvest. Springer, Cham, 2018, pp. 65-108
  36. 36. Agricultura. El cultivo de la papaya. [Accessed: 19 February 2021]
  37. 37. Ming R, Hou S, Feng Y, Yu Q , Dionne-Laporte A, Saw JH et al. The draft genome of the transgenic tropical fruit tree papaya (Carica papaya Linnaeus). Nature 2008; 452: 991-996
  38. 38. Cabrera-Ponce JL, Vegas-Garcia A, Herrera-Estrella L. Herbicide resistant transgenic papaya plants produced by an efficient particle bombardment transformation method. Plant Cell Rep 1995; 15: 1-7
  39. 39. Stepek G, Curtis RHC, Kerry BR, Shewry PR, Clark SJ, Lowe AE et al. Nematicidal effects of cysteine proteinases against sedentary plant parasitic nematodes. Parasitology 2007; 134: 1831-1838
  40. 40. Mansur F, Luoga W, Buttle DJ, Duce IR, Lowe A, Behnke JM. The anthelmintic efficacy of natural plant cysteine proteinases against two rodent cestodes Hymenolepis diminuta and Hymenolepis microstoma in vitro. Vet Parasitol 2014; 201: 48-58
  41. 41. Mansur F, Luoga W, Buttle DJ, Duce IR, Lowe A, Behnke JM. The anthelmintic efficacy of natural plant cysteine proteinases against the rat tapeworm Hymenolepis diminuta in vivo. J Helminthol 2016; 90: 284-293
  42. 42. Mansur F, Luoga W, Buttle DJ, Duce IR, Lowe AE, Behnke JM. The anthelmintic efficacy of natural plant cysteine proteinases against the equine tapeworm, Anoplocephala perfoliata in vitro. J Helminthol 2016; 90: 561-568
  43. 43. Mansur F, Luoga W, Buttle DJ, Duce IR, Lowe A, Behnke JM. The anthelmintic efficacy of natural plant cysteine proteinases against Hymenolepis microstoma in vivo. J Helminthol 2014; 760: 601-611
  44. 44. Berger J, Asenjo CF. Anthelmintic activity of crystalline papain. Science (80- ) 1940; 91: 387-388
  45. 45. Stepek G, Lowe AE, Buttle DJ, Duce IR, Behnke JM. In vitro and in vivo anthelmintic efficacy of plant cysteine proteinases against the rodent gastrointestinal nematode, Trichuris muris. Parasitology 2006; 132: 681-689
  46. 46. Stepek G, Buttle DJ, Duce IR, Behnke JM. Human gastrointestinal nematode infections: Are new control methods required? Int J Exp Pathol 2006; 87: 325-341
  47. 47. Stepek G, Lowe AE, Buttle DJ, Duce IR, Behnke JM. In vitro anthelmintic effects of cysteine proteinases from plants against intestinal helminths of rodents. J Helminthol 2007; 81: 353-360
  48. 48. Stepek G, Lowe AE, Buttle DJ, Duce IR, Behnke JM. The anthelmintic efficacy of plant-derived cysteine proteinases against the rodent gastrointestinal nematode, Heligmosomoides polygyrus, in vivo. Parasitology 2007; 134: 1409-1419
  49. 49. Stepek G, Lowe AE, Buttle DJ, Duce IR, Behnke JM. Anthelmintic action of plant cysteine proteinases against the rodent stomach nematode, Protospirura muricola, in vitro and in vivo. Parasitology 2007; 134: 103-112
  50. 50. Hounzangbe-Adote S, Fouraste I, Moutairou K, Hoste H. In vitro effects of four tropical plants on the activity and development of the parasitic nematode, Trichostrongylus colubriformis. J Helminthol 2005; 79: 29-33
  51. 51. Okeniyi JAO, Ogunlesi TA, Oyelami OA, Adeyemi LA. Effectiveness of dried Carica papaya seeds against human intestinal parasitosis: A pilot study. J Med Food 2007; 10: 194-196
  52. 52. Sciutto E, Hernández M, Díaz-Orea A, Cervantes J, Rosas-Salgado G, Morales J et al. Towards a practical and affordable oral papaya-based vaccine: A crucial tool for taeniasis cysticercosis control programs. In: Liong M-T (ed). Bioprocess Sciences and Technology. Nova Science Publishers, Inc., 2011, pp. 323-338
  53. 53. Betancourt MA, De Aluja AS, Sciutto E, Hernández M, Bobes RJ, Rosas G et al. Effective protection induced by three different versions of the porcine S3Pvac anticysticercosis vaccine against rabbit experimental Taenia pisiformis cysticercosis. Vaccine 2012; 30: 2760-2767
  54. 54. Hernández M, Jacquelynne G, Torres C, Luisa M, Gladis V, González F et al. Development of an Oral Anti-Cysticercosis Vaccine Delivered in Genetically Modified Embryogenic Callus of Papaya. Transgenic Plant J 4 2010
  55. 55. Chan HT, Daniell H. Plant-made oral vaccines against human infectious diseases-Are we there yet? Plant Biotechnol J 2015; 13: 1056-1070
  56. 56. Murashige T, Skoog F. A Revised Medium for Rapid Growth and Bio Assays with Tobacco Tissue Cultures. Physiol Plant 1962; 15: 473-497
  57. 57. Gamborg OL, Miller RA, Ojima K. Nutrient requirements of suspension cultures of soybean root cells. Exp Cell Res 1968; 50: 151-158
  58. 58. Widholm JM. The use of fluorescein diacetate and phenosafranine for determining viability of cultured plant cells. Biotech Histochem 1972; 47: 189-194
  59. 59. Pérez A. Establecimiento de células en suspensión de C. papaya que expresan el péptido KETc7 como propuesta de una vacuna contra la cysticercosis y la hemoncosis. Maestría en Biotecnología: Universidad Autónoma del Estado de Morelos. 2019
  60. 60. Caspeta L, Nieto I, Zamilpa A, Alvarez L, Quintero R, Villarreal ML. Solanum chrysotrichum hairy root cultures: Characterization, scale-up and production of five antifungal saponins for human use. Planta Med 2005; 71: 1084-1087

Written By

Marisela Hernández, Anabel Ortiz Caltempa, Jacquelynne Cervantes, Nelly Villalobos, Cynthia Guzmán, Gladis Fragoso, Edda Sciutto and María Luisa Villareal

Submitted: February 2nd, 2021 Reviewed: March 15th, 2021 Published: April 29th, 2021