**3***.* **Opportunities for Increased Income, Employment, and Food Supply in Seed Yam Production**

**3. Opportunities for Increased Income, Employment, and Food Supply in Seed Yam Production** Seed tubers are expensive, sometimes accounting for 63% of total variable production costs, and are bulky and expensive to transport (Manyong 2000; Agbaje et al. 2005). As planting material, the yam tuber has a meager multiplication ratio of 1:3 to 1:5 compared to some cereals (1:300). The low multiplication rate is a critical constraint to increasing yam production and productivity, resulting in the scarcity and high cost of quality seed yam (Aighewi et al. 2017). The consequence is that improved, released yam varieties are rarely found with farmers, and the use of farmer-saved seed is the norm. About a third of harvested yam tubers are reserved for seed for the next crop. Thus, from a West African production of 70.8 million tonnes, it is estimated that over 23.6 million tonnes, some of which would have been used as food, is reserved for planting the next crop (Figure 2). Continuous recycling of planting material often reduces quality by accumulating pests and diseases. After planting, some farmers can still reserve up to a third of the quantity of seed planted to replace those that would eventually not sprout due to poor quality (Aighewi 1998), further depleting a potential food and income source. Seed tubers are expensive, sometimes accounting for 63% of total variable production costs, and are bulky and expensive to transport (Manyong 2000; Agbaje et al. 2005). As planting material, the yam tuber has a meager multiplication ratio of 1:3 to 1:5 compared to some cereals (1:300). The low multiplication rate is a critical constraint to increasing yam production and productivity, resulting in the scarcity and high cost of quality seed yam (Aighewi et al. 2017). The consequence is that improved, released yam varieties are rarely found with farmers, and the use of farmer-saved seed is the norm. About a third of harvested yam tubers are reserved for seed for the next crop. Thus, from a West African production of 70.8 million tonnes, it is estimated that over 23.6 million tonnes, some of which would have been used as food, is reserved for planting the next crop (Figure 2). Continuous recycling of planting material often reduces quality by accumulating pests and diseases. After planting, some farmers can still reserve up to a third of the quantity of seed planted to replace those that would eventually not sprout due to poor quality (Aighewi 1998), further depleting a potential food and income source.

**Figure 2.** Yam tubers of up to 1 kg, which are good for local food and export, are used as seed and cut into minisetts of 30 to 50 g for multiplication. Source: Pictures by authors. **Figure 2.** Yam tubers of up to 1 kg, which are good for local food and export, are used as seed and cut into minisetts of 30 to 50 g for multiplication. Source: Pictures by authors.

The traditional methods to produce seed yam include cutting ware-sized tubers into seed size, harvesting the same crop twice (the first harvest is used for food and the second for seed), sorting seed size tubers, and using only the head portions of tubers (Aighewi et al. 2015). These options multiply yam very slowly with no guarantee of seed quality. This situation formed the basis for the Yam Improvement for Income and Food Security in West Africa (YIIFSWA) initiative of the International Institute of Tropical Agriculture (IITA). The focus was on developing methods to improve the quality of seed yam and rapidly multiply the seed to meet the needs of farmers for increased productivity. Any technology to rapidly multiply the crop is an opportunity to develop seed yam production-related businesses as a source of income, especially for youths and women. The development of hitherto The traditional methods to produce seed yam include cutting ware-sized tubers into seed size, harvesting the same crop twice (the first harvest is used for food and the second for seed), sorting seed size tubers, and using only the head portions of tubers (Aighewi et al. 2015). These options multiply yam very slowly with no guarantee of seed quality. This situation formed the basis for the Yam Improvement for Income and Food Security in West Africa (YIIFSWA) initiative of the International 178

4

non-existent seed yam enterprises will fill the gap in the supply of quality seed yam

and create job opportunities.

Institute of Tropical Agriculture (IITA). The focus was on developing methods to improve the quality of seed yam and rapidly multiply the seed to meet the needs of farmers for increased productivity. Any technology to rapidly multiply the crop is an opportunity to develop seed yam production-related businesses as a source of income, especially for youths and women. The development of hitherto non-existent seed yam enterprises will fill the gap in the supply of quality seed yam and create job opportunities.

#### **4. The Key Elements of the YIIFSWA Project**

YIIFSWA is a ten-year project, implemented from 2011 to 2021 with grants provided by the Bill and Melinda Gates Foundation (BMGF) to the International Institute of Tropical Agriculture (IITA) in two phases of five years each. The project's focus was to improve the productivity of yam through enhancing its seed systems.

#### *4.1. Summary of Achievement of the First Phase of the YIIFSWA Project*

The first five years of the YIIFSWA project (September 2011 to December 2016) facilitated activities to increase yam productivity (yield and net output) by 40% for 200,000 smallholder farmers in Ghana and Nigeria. Another goal was to generate international research goods that will double the income of 3 million yam producers in a 10-year horizon. During this first phase, YIIFSWA initiated the development of formal yam seed production systems, leading to the first set of certified seed yams for sale to ware yam producers in May 2016. This significant step in developing the yam seed market was critical to sustaining commercial production and marketing of high-quality seed yam. Diagnostics tools for virus detection and technologies for elimination of virus infected sources from seed production, high-ratio propagation technologies (HRPTs) such as the Temporary Immersion Bioreactor System (TIBS) and aeroponics, and seed yam quality management protocols as well as quality standards for seed yam certification were developed.

Over 65,000 farmers were trained to improve their seed production techniques using the adaptive yam minisett technique (YIIFSWA 2017), which is similar to what is used in the traditional seed production system (Aighewi et al. 2014). Yam-growing households in Ghana and Nigeria were characterized through a baseline survey (Mignouna et al. 2014a, 2014b). Economic assessments showed gains from the various technologies emanating from YIIFSWA and their relative profitability (Mignouna et al. 2020). Thousands of training materials (flyers, books, videos, and posters) were produced and disseminated to stakeholders to improve their capacities in seed and ware yam production, handling, and marketing (YIIFSWA 2017).

#### *4.2. Summary of Achievements of the Second Phase of the YIIFSWA Project (YIIFSWA-II)*

A primary goal of the YIIFSWA II project was to significantly increase yam productivity by at least 30% by delivering clean, quality seed yam of improved varieties to at least 320,000 smallholder farmers for long-term benefits in Nigeria and Ghana. This initiative aimed to improve the yield gains of the rural and urban poor, with more gender-equitable income for participants in the yam value chain. The project achieved its vision of demand creation, improving production systems and enhancing the enabling environment for improved varieties through four objectives:

i. Increase productivity and income by empowering smallholder ware yam producers with the seed of improved varieties. Three improved and released varieties of *D. rotundata* (TDr 89/02665 named Asiedu and TDr 95/19177 named Kpamyo) and one of *D. alata,* (TDa 98/01176 named Swaswa) were introduced to farmers in demonstration plots to show the potential of improved varieties. Their mean productivity was higher than that of the local farmers' preferred varieties amounting to 38% productivity increase.

The project developed functional and sustainable seed systems by strengthening national agricultural research institutions to produce high-quality breeder seed yam of improved varieties using the TIBS. In 2020, the Ghana Crops Research Institute (CRI), with the assistance of the YIIFSWA-II project, produced 350% of its target of 21,840. The Savannah Agricultural Research Institute (SARI) in northern Ghana, with the support of the project, established its first TC Laboratory since inception and produced 399% of its yearly target of 8736 breeder seed plantlets using TIBS. The National Root Crops Research Institute (NRCRI- Umudike) of Nigeria produced 104% above its target of 21,840 breeder seeds in 2020. The breeder seeds produced by NRCRI were partially distributed to two private seed companies (Nwabudo Agro Seeds and Inputs Ltd. and Strategic Seeds Nigeria Ltd.) and to the Federal Ministry of Agriculture and Rural Development. In total, 221,735 breeder seeds were produced by IITA and NARIs within the year 2020 exceeding the target production of 71,872 by 309%.

ii. YIIFSWA developed, demonstrated, and trained private seed companies on foundation seed production models using best management practices in aeroponic and hydroponic systems. These activities were aimed to deliver high-quality seeds of improved varieties to farmers at the right time and prices to encourage adoption through commercial seed entrepreneurs. One of the primary outcomes of the project was to establish a functional, scalable, and sustainable foundation and commercial seed systems that are driven by the private sector. The project accomplished this by identifying and developing the capacities of nine private seed companies in Ghana (3) and Nigeria (6). These seed companies together with some of the NARIs produced about 3.6 million foundation tubers of 25 g each in 2020.


The implementation of YIIFSWA II ensured that all the advances made in establishing the yam seed system by YIIFSWA were fully transferred to and operationalized by National Agricultural Research Institutes (NARIs) and private seed companies. Additionally, enhanced on-farm agronomic packages were accessible and utilized by seed and ware yam farmers to increase productivity and income generation. Beneficiaries of the project include seed and ware yam producers and their family members, processors, marketers, transporters, and consumers.

#### **5. Novel Yam High Ratio Propagation Systems: Outcomes and Prospects in Adaptation to Climate Change and Nutrition Security**

The YIIFSWA project developed and standardized a tissue-culture-based heat therapy combined with a meristem culture procedure to generate virus-free yam planting material. This procedure had a 73% success rate in eliminating the yam mosaic virus (YMV), the most frequently contaminating virus in yams in West Africa (Balogun et al. 2017a). This procedure established YMV-free stocks of 25 yam genotypes consisting of improved varieties and landraces, which were used as stocks for rapid bulking of clean planting material using the TIBS, an advanced high-ratio in vitro propagation technology also standardized by YIIFSWA.

Plantlets from TIBS are of better quality than those from conventional tissue culture (CTC). TIBS plantlets are more vigorous and resilient to post-flask acclimatization. Due to more efficient process control, large batches are handled more easily for scale-up propagation with lower risks of mix-ups. The propagation ratio in TIBS was five to six per plantlet every eight to ten weeks compared to three to four every 12 to 16 weeks in CTC (Balogun et al. 2017b). Up to 300 single-node vine cuttings were obtained per plant that was derived from TIBS plantlet and grown aeroponically (Maroya et al. 2014c, 2017), while the drip system hydroponics further saves on electricity needs in the production system (Balogun et al. 2020, 2021). About 100 single-node vine cuttings can be made per TIBS plantlet using hydroponics after eight weeks of growth, with 95% rooting success followed by field planting. After five months of hydroponic growth, the seed tubers ranged from 5 g to 220 g per plant.

Aeroponics yam propagation started under the YIIFSWA project in 2013 at IITA-Ibadan, Nigeria (Maroya et al. 2014a, 2014c). Atomizing nozzles ensure the most effective nutrient delivery in this system by turning the nutrient solution into a mist, which is absorbed through the cell walls of the plant's roots by osmosis. Genotypes of both *D. rotundata* and *D. alata* were successfully propagated with aeroponics using both pre-rooted and freshly cut two-node vine cuttings. Three types of planting materials are generated from the aeroponics production system: mini-tubers which range from 0.2 g to 110 g depending on the genotype, harvest age, and the composition of the nutrient solution, the aerial bulbils from both water yam and white yam varieties, and single-node cuttings from vines (Maroya et al. 2014b). All the propagules had a propagation rate of over 90%. Yam vine cuttings were previously known to grow slowly, but those from aeroponic plants produced new leaves 14 to 21 days after planting. The single-node vine cuttings from aeroponic plants produce an average of 200 to 300 cuttings per plant in four to six months. A manual was produced to help private seed companies to establish their aeroponics

systems with step-by-step instructions for building and managing the system for foundation seed production (Maroya et al. 2017). In 2017, the YIIFSWA-II project funded each of five selected seed companies with USD 30,000, half of the cost to build a screen-house and an aeroponics system. The project produced and distributed virus-indexed plantlets for multiplication to seed companies. Relevant stakeholders were trained, including postgraduate degree students, to apply various propagation techniques through participatory research. After training, new technical information on the high ratio propagation of yam was constantly provided, accompanied by backstopping partners and other stakeholders.

The capacities of the seed regulatory agencies of Nigeria, the National Agricultural Seed Council (NASC); and of Ghana, the Plant Protection and Regulatory Services Directorate (PPRSD); were also enhanced. As coordinators in the management of seed quality in their respective countries, they were trained and provided with equipment to ensure that high-quality seeds of authentic varieties were produced and sold.

#### **6. Improved Yam Seed Systems as Key to Building Resilience in Production to Enhance and Sustain Food Security**

A functional yam seed system is essential to build resilience in the ware yam production systems. As the most critical and expensive input of yam production, the seed should be healthy to generate healthy plants in the field and reduce yam's current high postharvest losses estimated at 30%. Seed health influences the shelf life of harvested ware tubers or postharvest storage quality since pests and diseases are easily transferred from the field into storage and over different seasons through the seed. Therefore, a yam seed system must have protocols to ensure the production of seed tubers at a high rate of multiplication. Rapid propagation of pest- and disease-free seeds and a network of commercial seed producers within a formal seed system are crucial for disseminating improved varieties for sustained and increased productivity. Extension services were used to educate farmers on plant health management in the field to recognize pest and disease symptoms and carry out positive selection practices to sustain good yields.

Technologies for breeder seed production were transferred to the following NARIs, the National Center for Genetic Resources and Biotechnology (NACGRAB) and the National Root Crops Research Institute (NRCRI) in Nigeria, and the Crops Research Institute (CRI) and Savanna Agricultural Research Institute (SARI) in Ghana. These institutions were provided with equipment and supplies for backup electricity, TIBS, post-flask handling and documentation, and initial clean stock of planting

materials. The NARIs now provide clean breeder planting materials to private seed companies for foundation seed production.

The innovations developed by YIIFSWA to improve yam seed systems have added value to the yam crop by using vines for propagation. Before now, yam vines were mainly considered as the apparatus for manufacturing photosynthates for storage in tubers. In the new seed production systems, leafless nodal cuttings are used in TIBS, while those with leaves are planted in various hydroponic systems or rooted and planted in the field (Figure 3). The nodal cuttings with leaf can also be planted directly in the field for seed yam production. Methods of producing minitubers of less than 10 g, which perform excellently in seed yam production (Aighewi et al. 2021), have also been developed. If widely adopted by seed companies, there would be less pressure on using yam tubers for food and seed. Six seed companies in Nigeria and three in Ghana are using the innovations developed and scaled by the YIIFSWA project for seed production.

**Figure 3.** Types of yam planting materials (**a**). In vitro single node cuttings for multiplication in temporary immersion bioreactor systems; (**b**) Ex vitro nodal cuttings with leaf for multiplication in various hydroponics systems, soil, and composite media; (**c**). Minitubers from ex vitro single node plants for vine and tuber planting material production in the field or screenhouse. Source: Pictures by authors.

The HRPTs developed by YIIFSWA are contributing to the efficient production of end-user preferred improved varieties. According to previous studies by CIRAD and partners, flour from dried chips made from the late maturing, multiple tubering (5–10 tubers per plant) varieties of *D. cayenensis* and *D. rotundata* with relatively high dry matter content (around 40%) and tolerance to poor soil conditions were most demanded for preparation of *amala,* a popular food in Nigeria and Benin. This link between food demand and preferred variety confirms that seed systems need to choose their promoted varieties based on market demand. However, the delivery rate of improved varieties is slow due to the low propagation ratio in traditional yam cultivation, where cropping is seasonal, with a single annual production cycle that is significantly limited by season (Orkwor and Asadu 1998). Weather elements are also crucial for yam production ranging from water, light, nutrients, and temperature. Yam requires 20 ◦C to 30 ◦C for optimum growth. However, according to the IITA's Geospatial laboratory, in 2015, temperatures lower than 20 ◦C were recorded only in January and December, a period that is typically outside the crop growth period, while at least 7 out of the 12 months in 2005, 2010, 2015 and 2017 recorded higher than 30 ◦C (Personal communication) with associated rising air temperature and carbon dioxide levels. The impact of climate change on the yam yield, including its vulnerability to climate-change-related soil conditions, has been discussed by Srivastava et al. (2012).

Just as drought causes significant losses, floods also cause significant yield losses (Balogun and Gueye 2013). These climate-related scenarios force farmers to vacate flood- or drought-prone farmlands or risk immense losses due to inadequate photosynthetic rates, tuber bulking, and premature senescence. While there have been significant breakthroughs in the real-time availability of geographical information systems, its utilization in timing yam production is yet to be entirely scaled and adopted at the level of producers and processors. The scarcity of clean planting materials (Aighewi et al. 2015, 2021), dormancy of the tuber for about four months, and uncontrolled sprouting after dormancy break limits the control of the cropping calendar by farmers and reduces profits (Craufurd et al. 2001). This established need for resilient, climate-smart mitigation strategies is also addressed by the gains from the YIIFSWA project, especially in terms of yam cropping cycles. Seed production can be carried out in controlled environments during the offseason, and ware yam production timed to maximize the available rainy period.

Table 1 shows the timing of seed yam production based on the traditional system relative to the HRPTs. Two cropping cycles are possible per year, from May to December and October to August. Previously, 14 t/ha at 40,000 stands for seed yam production was recorded. If the same land is used, up to 30 t/ha will be produced per cycle when vine cuttings are used, giving 60 t/Ha per year. With continuous availability of seed yam from the HRPTs, ware yam can be produced throughout the year, combining the traditional cycle of October to July with the HRPT cycle of May to December. This scheme is more amenable to delayed rains if supplemental irrigation is provided. Significantly more land can also be used for yam production due to 500-fold more seed yam, which culminates in more food availability.


**Table 1.** Seed yam production based on novel high ratio propagation technologies.

AS = aeroponic system; HS = hydroponic system. Source: Maroya et al. (2022).

van Etten et al. (2017) has established that the access farmers have to quality seed and the functionality of seed systems in relation to production, distribution, innovation, and regulation determines the efficacy in contributing to sustainable agrobiodiversity and food systems. Available evidence has shown that the YIIFSWA II project has impacted all four aspects. However, it is necessary to incorporate good practices for efficient soil and water management as well as good agronomic and agroecological practices that mitigate and enhance the adaptation to climate change in future projects.

An additional benefit of the novel high-ratio propagation system is the mitigation against hidden hunger. Nutrients can be dosed to yam grown in aeroponics and hydroponics systems. High-quality planting materials of improved varieties are disseminated as reported by a study conducted by YIIFSWA (unpublished) to determine the nutrient components that limit the propagation ratio and quality of plantlets in the TIBS. Using white and water yam varieties, Kpamyo and Swaswa, respectively, the nitrogen (N), potassium (K), and phosphorus (P) concentrations in culture medium were determined every two weeks for ten weeks and in plantlets at transplanting. After eight weeks of culture, the pH and refractometer values for medium acidity and sugar concentration were determined in the varieties. In Swaswa, N, P, and K in the medium reduced by 83.8, 96.2, and 28.7%, while it was 63.3, 61.2, and 23.1% in Kpamyo over the ten-week period. Reduction in K concentration at two weeks was limited to Swaswa. In Kpamyo, adding P at two weeks and N at six weeks would be beneficial. Thus, it is necessary to consider diet requirements from seed production to determine optimum fertigation regimes. This will ensure that plants absorb the available nutrients in sufficient quantities for storage in tubers, which can also be fortified with additional micronutrients if processed into other products after harvest.

#### **7. Outcomes and Impact on Reducing Hunger by Raising Food Security**

Food security is commonly defined as the access by all people to sufficient food for active and healthy life (World Bank 1986). Three critical dimensions implicit in this definition are (i) the availability of sufficient quantity and appropriate quality of food supplied through own production or otherwise; (ii) access by all households and individuals to enough and adequate resources to acquire such food; and (iii) the utilization of this food through an adequate diet, water, sanitation, and health care (Timmer 2012). In the developing world, household food security is largely linked to food availability from household production. Gifts and transfers from friends and relatives also play essential roles. Food purchased are also common but limited due to a lack of liquidity.

#### *7.1. Changes Perceived in Household Food Consumption Status of Yam Growing Areas of Nigeria and Ghana*

Food deficit was the main periodic shock experienced by most households across the yam-growing areas of Nigeria and Ghana. This type of vulnerability results from qualitative analysis considering the respondents' perception about the number of households affected by food shortages and the frequency of food shortages during a

season. To assess the farm household's food consumption during surveys organized by the YIIFSWA project, memory recall on different food shortage scenarios in the past 12 months was employed.

Following the baseline survey, an end-line survey was performed within the same major yam-producing zones in Nigeria and Ghana using the same multistage, random sampling design, drawing on 1400 households based on the same sampling frame (Mignouna et al. 2016). The respondents were asked whether their households had sufficient food during the previous year. Figure 4 shows how households perceived their food security status. This perception was observed to be different between the two rounds of surveys. After the project's first phase, changes in shocks faced by households pursuing their livelihood strategy were noticed by comparing their baseline situation to the end line.

**Figure 4.** Changes in household food consumption status in a 12-month period for yam growing households in Nigeria and Ghana. Source: (Mignouna et al. 2014a, 2014b, 2017a, 2017b).

In Nigeria, from the baseline to end line, households reported a reduction in food shortages throughout the year from about 5% to 1%. In comparison, reports on occasional food shortages decreased from 32% to about 26% (Figure 4). Households that reported no food shortage, but no surplus increased from about 44% to 54% during the two periods. Households reported that food surplus was almost unchanged during the same periods. In Ghana, the remarkable observations

from baseline to the end line are that the proportion of households that reported occasional food shortage decreased by 27% points from about 62% to 35%, while the households that reported having a food surplus increased by 26% from about 8% to 34% (Figure 4). Almost no difference was observed between households that reported no food shortage but no surplus and food shortage throughout the year.

Positive changes in food consumption status were reported, and these probably resulted from increased productivity due to interventions of the YIIFSWA project. They are good indicators of food security improvement in the region, likely due to YIIFSWA's interventions. This translates into a contribution of the project in reducing vulnerability to food insecurity among rural households in the surveyed areas.

#### *7.2. Project Impact on Food Security*

Total household expenditure included expenditures on non-food items and consumables. Under food expenditure, all the food items consumed by the household during a year were collected. Food consumption included food that the household purchased, produced, and received from other sources. The total expenditure on food, obtained by aggregating expenditure on all food items, was used to estimate the project's impact on food security. The total expenditure for each food group was calculated by aggregating the expenditure of all food and non-food items falling within a group. The results of the propensity score matching (PSM) presented in Table 2 show that the Average Treatment Effect on the Treated (ATT) of YIIFSWA project on food security (per capita expenditure on food) of beneficiaries in Nigeria and Ghana were about USD 55 and USD 94 for the Radius Matching technique. These results imply that the beneficiaries that profited directly or indirectly from the project are more food secure than the non-beneficiary farmers.

In summary, this study sought to find out the average treatment effect on the treated, which gives the average effect of the project on food security. The results showed a positive impact on food security, implying that the increase in productivity generated by the project interventions led to increased household food security and poverty reduction in the region.


#### **Table 2.** Impact of YIIFSWA on food security.

ATT: Average Treatment Effect on the Treated; ATU: Average Treatment on Untreated; ATE: Average Treatment Effect. USD 1 = GHS 1.85 = NGN 157. Source: (Mignouna et al. 2014a, 2014b, 2017a, 2017b).

#### **8. Conclusions**

The YIIFSWA initiative has focused on efficient seed yam propagation and production, significantly impacting the yam value chain. The project has developed high-ratio propagation technologies and seed-health management innovations to improve the seed quality and increase productivity by supplying yam farmers with healthy seeds. The new methods emphasize using vines and minitubers of less than 10 g to produce seed tubers and free large quantities of tubers for food. Stakeholders' capacities were enhanced through training on improved seed production methods and equipment to build resilience and sustainability, and increase seed production for income, nutrition, and food security.

#### **9. Recommendations**

The gains from the innovations contributed by the YIIFSWA project are visible. However, greater emphasis is required for scaling innovations, especially in certified seed production, for higher impact. Further research efforts are necessary to identify cheaper alternatives in the novel seed yam propagation technologies that will increase the profit margins in seed yam businesses. Studies are also required to identify the most appropriate agronomic practices and control of yam tuber dormancy to allow integration into cropping cycles in the face of production intensification and the changing climate.

**Author Contributions:** Conceptualization, B.A.; writing—original draft preparation, B.A., R.A.; writing—review and editing, B.A., N.M., R.A., D.M., M.B., P.L.K.; project administration, N.M., R.A.; funding acquisition, IITA: R.A., N.M., P.L.K., M.B., D.M. and B.A. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research was funded by the Bill and Melinda Gates Foundation, bearing Investment Number OPP1159088. Grant to the International Institute of Tropical Agriculture, Ibadan, Nigeria, November 2016 to 31 December 2021. IITA receives funding for research on yams from the CGIAR Research Program on Roots, Tubers, and Bananas (CRP-RTB) between 2011–2021 and the ongoing CGIAR Seed Equal Initiative, both supported by CGIAR Trust Fund donors.

**Conflicts of Interest:** The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

#### **References**


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