Soybean [Glycine max (L.) Merr.] is an important leguminous crop, widely grown as food and feed for man and livestock. It is also grown for its oil, nitrogen-fixing ability, and fuel production as biodiesel (Kofsky et al. 2018; Nawaz et al. 2017). The sap-sucking hemipteran insect pests from the family Aphididae, especially soybean aphid (SA, Aphis glycines), pea aphid (PA, Acyrthosiphon pisum), and foxglove aphid (FA, Aulacorthum solani Kaltenbach), are constraints to soybean production (Blackman et al. 2000; Nagano 2001; Song et al. 2019). Global climate change causes temperature fluctuation which enhances overwintering survival time, the generation number of FA, and the population spread of these aphids due to an altered ecology and aphid habitat (Skendzic et al. 2021). High fecundity of these aphids under optimal conditions supports a rapid population growth rate making their control difficult. Although soybean plants have provided a resistance mechanism to hinder the herbivory effect of aphids, aphids overcame it (Neupane et al. 2019). The impact of these aphids resulted in heavy reliance on insecticide application (Ono 2006) which increases the cost of soybean production (Steffey 2004) with a potential insecticide resistance effect. To overcome this bottleneck, developing insect pest resistance soybean plants is an economic and efficient strategy (Lee 2015).

FA insect pest activities in Korea, and Japan, threaten soybean production. Japan recorded a 90% reduction in soybean yield due to the infestation of the FA in 2000 (Nagano 2001) and its soybean pest occurrence was also identified in Korea (Kim 1991). It has a wide host range of about 540 plant species (Jandricic 2010) including, ornamentals (Blackman et al. 2000; Jandricic 2010), and common weeds (Capinera 2001). Furthermore, this aphid is considered a vector of about 45 plant viruses (Jandricic 2010; Miller 1997; Yovkova et al. 2013) causing further damage to plants. FA feeds on soybean plants by sucking out the sap from phloem tissues with its stylet and the release of toxic saliva that deforms both leaves and fruits (Sanchez et al. 2007). The feeding habit adversely affects soybean with leaf deformation, yellowing of leaf veins, sometimes necrosis, and plant wilting. A considerable amount of honeydew is excreted by FA which enhances mold growth on soybean leaves with a decrease in photosynthetic activities (Morkunas et al. 2011). The degree of damage depends on the FA density on the soybean.

To date, several soybean cultivars have been identified as resistant to FA. The first study on resistance in soybean to FA started in 2008 (Kim 2008), and two Raso genes, Raso1 from cultivar ‘Adams’ (Ohnishi et al. 2012) and Raso2 from soybean variety ‘PI 366121’ (Lee 2015), were identified and fine-mapped on chromosomes 3 and 7, respectively. Therefore, studies to understand the inheritance of FA resistance are ongoing as means to develop resistant cultivars through a breeding program. The main objectives of this review were i) to provide summarized information about the biology of FA, ii) to highlight knowledge on soybean resistance to FA, and iii) to suggest future challenges of research on soybean-FA interaction and breeding strategies for effective management of FA in soybean production.

Origin and distribution of Foxglove Aphid

FA also referred to as glasshouse potato aphid is a known as insect pest of different crops. It is believed to be a native of Europe (Blackman et al. 1984) but is now found worldwide especially in temperate and Mediterranean climatic regions as a crop pest hence, regarded as cosmopolitan (Essig 1942; Michelangelo 2019). It was originally described by studying its activities on its primary host which are common foxglove Digitalis purpurea L.- reasons for the coined name, foxglove aphid (Patch 1928), common perennial hawkweed Hieracium spp (Wave 1965), and potato Solanum tuberosum (Blackman et al. 1984) in North America. The emergence of the FA as an insect pest is an indication of the impact of climate change (Skendzic et al. 2021). A few decades back, soybean insect pests were lepidopteran and coleopteran. But global warming effected a change of pest’s type to stink bugs and sap-sucking hemipteran aphids, such as FA (Bansal 2013). FA mouthpart stylet is well adapted for piercing and sucking to extract phloem sap from the host plant. Genetic analysis studies on FA reveal that they are a species with varying biotypes (Miller 2009). Previous research shows that FA infestations had a devastating effect on soybean in Asia (Kim 1991; Nagano 2001), but less destructive in North America. These results suggest the presence of various biotypes, which is a unique phenomenon with aphid species (Miller 2009). The genetic diversity of A. solani within the East Asia population sampled differs based on geographical location. Apart from its wide host range, the biological variability of the FA population globally remains largely uncharacterized. Information on the biology and ecology of FA is still limited.

Biology of FA

FA is a shiny yellow, green, or yellowish-green insect pest (Capinera 2001) having on the abdomen dark-green patches around the base of their short cornicles (Fig. 1A). Aphid has long legs and antenna which are distinctively marked with black banding as shown in Figs. 1B and 1C (Stoetzel 1994). Adult oval body size is around 1.8 - 3.0 mm (Stoetzel 1994). In the process of feeding on the host plant by sucking its sap, plant vigor gradually diminishes in Figs. 1C, 1E and 1F. Under the lower temperatures, FA possesses a high reproductive rate ability which can support exceeding natural enemy suppression capacity (Alotaibi 2008). Therefore, biological control agents are incompetent under low temperatures in the management of FA (Bellefeuille 2021). FA has a complex but short lifecycle which enhances rapid population build-up. Aphid undergoes two forms of lifecycle (holocyclic and anholocyclic) on a host plant. Reproduction could be either sexual or asexual, even though asexual reproduction is most common. Eggs of FA survive winter season and wait till spring season before hatching into many nymphs. The nymphs immediately start to feed on plant sap and grow rapidly. FA has five nymphal instars (Capinera 2001) and undergoes molting four times before attaining the adult stage. At the end of each molt, the aphid leaves behind its shed skin on the host crop. The emerging adult aphid can actively reproduce for varying days depending on prevailing temperature while the females produce an average of 60 (25-81) nymphs (Capinera 2001). Adult aphids can either be winged (alatae) or wingless (apterae). Interaction between FAs and their environment is greatly affected by the effect of global warming (i.e., temperature) on its physiological function and life cycle (Hulle et al. 2010).

Figure 1. Morphology of foxglove aphis and its damage in soybean leaves. Dorsal (A) and ventral (B) view of foxglove aphid, feeding on the host plant by sap-sucking (C), and foxglove aphids on soybean stem (D). Soybean leaf damage by foxglove aphids in Glycine max (E) and Glycine soja (F).

Factors affecting foxglove aphid development

The development of FA is directly affected by prevailing temperature and host plant nutritional content (Seo et al. 2020). The mortality of nymphs was examined at various temperature and type of host plants (Kikuchi 2018; Kim 1991). The results showed that nymphs of FA showed 100% mortality at 30°C, and numbers of nymphs produced by adult females of FA was higher at 20°C than 28°C. Furthermore, nymphal stage duration showed significant difference based on the host plants, and temperature (Jandricic 2010). In temperate regions of the world, FA is found to survive in the winter season due to its adaptability at a lower developmental threshold (LDT). The aphid adapts underestimated LDT values between 3°C - 5°C on different crops (Jandricic 2010; Seo et al. 2020). For example, during the winter in Scotland, an anholocyclic form of FA could survive on weeds. A temperature range of 12.5 to 25°C supports high fecundity in the aphid, which could lead to outbreak (Kim 1991). In soybean, FAs exhibited the highest mean daily fecundity at 25°C and 20°C with cumulative A. solani (ca.) 3.6 and 3.8 nymphs daily produced respectively per adult(Seo et al. 2020). Nutrient source (i.e., host plant leaves or other organs) also affected the fecundity. FAs tend to reproduce only few nymphs, when they were fed with host plant leaves with inadequate or excess soluble nitrogen, due to impaired reproduction (Atlihan et al. 2017; Awmack et al. 2002; Chen et al. 2018).

Identification and utilization of soybean accessions with resistance to FA

Plants have their unique immune system. It naturally has evolved resistance genes (R genes) that can recognize the presence of insect pests (Kourelis et al. 2018) with a possible prompt in response to hinder pest invading capacity to feed (Natukunda et al. 2019). A successful plant breeding program requires a large germplasm collection with maximum genetic diversity (Chheda 1982). Crop cultivars are known to possess low genetic variation in resistance related traits while wild crops have a wide genetic variation in resistance-related traits. Therefore, increasing crop species diversity is needed to tackle pest damage (Dainese et al. 2019; Isbell 2017). Soybean accessions in Japan and Korea have been subjected to susceptibility tests for FA to identify susceptible and resistant cultivars to FA. Currently, a very little portion of the accessions has been identified to be resistant (Table 1).

Table 1 . Identified soybean genes that confer resistance against FA.

S/N	Soybean Cultivar	Origin	Description	Reference
1	IT 104704	Korea	Strong antibiosis effect against aphid	(Koh et al. 2020)
2	IT 188399	Korea	Strong antibiosis effect against aphid	(Koh et al. 2020; Lee et al. 2008)
3	PI 230977		Has strong aphid resistance	(Koh et al. 2018; Koh et al. 2020)
4	PI 366121	Japan	Showed antixenosis and antibiosis resistance against aphid	(Kim et al. 2021; Koh et al. 2018; Koh et al. 2020; Lee 2015)
5	PI 548502	Japan	A high degree of aphid antibiosis resistance	(Ohnishi et al. 2012; Takahashi et al. 2002)
6	Tohoku149	Japan	Has strong aphid resistance	(Sato et al. 2013; Sato et al. 2014)

Host plant resistance is the heritable trait a plant species possess that enhances a decrease in insect pest population and activities on such plants (Dogimont et al. 2010). There are three categories of host plant resistance, antibiosis, antixenosis, and tolerance. Antibiosis could suppress the growth and development of insect pest. Antixenosis is a non-preference reaction of insect pest to host plants, caused by the morphological or chemical factors of host plant, which affect the insect pest behavior (Kamphuis et al. 2013; Smith 2005; Smith et al. 2012). In soybean, FA resistance responses are antixenosis and antibiosis (Koh et al. 2018; Lee 2015). Host plant aphid resistance is a promising control strategy with the advantage of cost-effectiveness, environmental safety, and compatibility with other control strategies.

In the case of SA, several resistant soybean cultivars were identified with different resistance to Aphis glycines (Rag) genes (Hesler et al. 2021) on soybean chromosomes 7, 13, 16, whereas only a few resistant soybean cultivars have been identified to possess resistance to Aulacorthum solani (Raso) gene. Some soybean cultivars were identified to exhibit antixenosis resistance effect against PA (Stec et al. 2021) due to the presence of flavonoids (genistein, kaempferol) in leaf tissues although such studies on the effects of bioactive compounds in leaves of identified cultivars resistant to FA has not been carried out.

The identified FA resistant cultivars were Adams and wild soybean Glycine soja PI 366121 with Raso1 and Raso2 genes on soybean chromosomes 3 and 7 respectively (Table 2). Also, soybean cultivar Tohoku 149 expressed resistance against FA having a unique profile of two metabolites which are trigonelline and S-methylmethionine. To avoid yield losses associated with FAs, securing resistant genetic resources is required.

Table 2 . Identified soybean genes that confer resistance against FA.

Gene	Chromosome	Soybean Cultivar	Form of Resistance	FA Biotype	References
Raso1	3	PI 548502	Antibiosis resistance	Japanese	(Kamiya 2008; Ohnishi et al. 2012)
Raso2	7	PI 366121	Antixenosis and Antibiosis resistance	Korea	(Kim et al. 2021; Lee 2015)

Most of the identified resistant cultivars are yet to be genetically and molecularly characterized. Soybean wild germplasms possess independently evolving genotypes within a geographical location with the prospect of improving soybean genetics against FA. Identifying wild soybean relatives with FA resistance trait is ongoing with the view of transferring the trait to soybean cultivar. Presently, characterized soybean germplasm includes Plant Introduction PI 366121 and Glycine soja wild soybean (Table 1 and Table 2) which showed resistance to FA, The PI has been successfully crossed with Williams 82, a susceptible soybean cultivar to FA (Kim et al. 2021; Lee 2015).

Novel QTL genes for FA Resistance in Soybean

The Raso1 gene was mapped on Adams or Shokukei-10 soybean on chromosome 3 within a 3.7 centiMorgan (cM) region between the simple sequence repeat (SSR) markers Satt009 and Satt530 (Ohnishi et al. 2012) which is a different location from the Rag1 gene in soybean Dowling cultivar mapped to a 12 cM region on soybean chromosome 7 between Satt435 and Satt463 (Li 2007) and other Rag genes.

In fine-mapping Raso1 on chromosome 3 at a 63-kb interval between markers Gm03-11 and Gm03-12 of the Williams 82 genome sequence, three candidate genes, which are Glyma03g04530, Glyma03g04500, and Glyma03g04510 were found through Phytozome v7.0 (Ohnishi et al. 2012). The Glyma03g04530 gene is a typical type of resistant R gene with is made up of the NB-ARC domain and a leucine-rich re-peat-containing gene (NBS-LRR).

Resistance gene Raso2 was detected in a major QTL region on chromosome 7 and several minor QTLs. Studies on the QTL on chromosome 7 show a negative additive effect value which suggests the possibility of the effect originating from PI 36612 resistant parent. The lower value of the log of odds (LOD) and phenotypic variance explained (PVE) % for QTLs on chromosomes 3, 6, and 18 suggested they are minor QTLs while higher values of LOD and PVE % on chromosomes 7 imply it is a major resistance QTL for antibiosis and antixenosis (Lee 2015). Therefore, the increased resistance against FA in PI 366122 is mainly due to this major QTL located in chromosome 7. The location of these major QTL is between SNP markers BARC-042815-08424 and BARC-015945-02020 at regions 71 cM and 84 cM which is 13 cM apart on chromosome 7.

Mapping of the Raso2 gene was earlier carried out using 414 SNPs out of the 1,536 SNP loci of the GoldenGate® assay (Lee 2015). Further mapping of the Raso2 gene used the same population with 28,752 SNPs of the 180K Axiom^® SoyaSNP array (Kim et al. 2021) could narrow down average distance value of 0.15 cM among SNP markers mapped compared to the previous study value of 6.8 cM that covered the entire genomic regions of soybean. During linkage mapping construction, a narrowing down of Raso2 interval was carried out from 2.2 Mbp (13 cM) of 275 annotated genes to 76 Kb (1 cM) of 8 annotated gene models using the 180K Axiom ^® SoyaSNP assay. On chromosome 7 of soybean varieties PI 366121, PI 587732, and Dowling, different resistance genes for FA and SA respectively, were mapped on different physical positions. The positioning of Raso2 in PI 366121 is 7 Mb while SA resistance genes were mapped in PI 587732 and Dowling on 43 Mb regions of chromosome 7.

Aphids are generally known for their peculiar ability to swiftly adapt to new varieties of plants that earlier express resistance. This complicated condition was regarded as an “arms race” between aphids and plants (Botha 2013). The challenges from soybean aphid resistance development in overcoming soybean resistance, therefore, suggested the stacking of various Rag genes to generate reliable resistance as a solution. Equally, the process of breeding soybean against FA is quite complicated due to the possible genetic variability of the FA population. There are differences in the ways FA biotypes attack the varying soybean cultivars. The presence of different FA biotypes recently threatens the process of breeding against a FA biotype. A soybean cultivar with resistance ability to a FA biotype may not be able to resist other FA biotypes. For instance, resistance against the Japanese biotype FA by Adams and Tohoku 149 does not confer resistance against the Korean biotype FA (Lee 2015). That is, the effectiveness of a resistant soybean cultivar to a FA biotype across different geographical locations is not certain. Therefore, it is a concern for soybean breeders on the development of soybean cultivars with the ability to universally resist every available FA biotype. Though breeding of resistant soybean cultivars against FA is promising with management of FA, a new FA biotype could possibly break resistance in soybean cultivars in the future. For instance, in Ohio USA, a new biotype overcame the Rag1 resistance gene for soybean aphids (Kim 2008). This limitation in the use of resistance genes in resistant cultivars could be possibly solved using gene pyramiding. It involves combining R genes in resistant cultivars to produce plants with active defense responses across different biotypes of insects (Natukunda 2021). Though the molecular mechanism remains unclear, the pyramiding of varying resistance genes into single plant genotype is known to confer enhanced resistance on the selected cultivar at the phenotype level.

With the possibility of FA overcoming resistance in the few identified resistant soybean cultivars, there is a need to identify more soybean genetic resources with inherent resistance. This involves the screening of large-scale soybean germplasm which will be assayed for their performance after infestation by FA. Then molecular and genetic characterization of the identified resistant variety for novel resistance gene is carried out. The process involves crossing, making genetic populations (F₂, recombinant inbred line (RIL)), genotyping, phenotyping, and QTL analysis. Little attention is given to genetic and molecular information on soybean resistance to FA when compared to soybean resistance to SA.

The discovery of new resistant genes against FA can be followed by combining its effect with the already reported resistance gene for sustaining and durable host plant resistance. A good understanding of the molecular mechanisms that support enhanced resistance in pyramiding genes would facilitate more directed approaches for crop improvement. Further studies using proteome and transcriptome analysis on the identified Raso gene are required. Since the dynamic interplay of molecular and physiological responses of soybean after infestation by FA is not fully understood, further investigation is still required (Enders et al. 2021). It is important to assess the bioactive compound produced in leaves of identified cultivars resistant to FA through molecular studies. Plant hormones such as salicylic acid, gibberellic acid, ethylene, abscisic acid, and jasmonic acid, are related to some metabolic pathways that regulate plant-aphid interactions. Therefore, studies on the impact of phytohormone biosynthesis in FA soybean-resistant cultivars should be carried out.

In genome sequencing research, unlike soybean aphids and pea aphids (464.3 Mbp and 302.9 Mbp), genomic information of the FA has not yet been revealed (Giordano et al. 2020; International Aphid Genomics 2010; Nouhaud et al. 2018; Wenger et al. 2020). Therefore, the study of whole-genome sequence assembly of FA is needed to understand FA adaptation to soybean, diversity of FA, and biotype evolution. Genetically characterizing FA biotypes will explain the mechanisms of FA virulence on resistant soybean cultivars and hence, explore the use of host plant resistance to achieve optimum soybean production.

In conclusion, the response of a few identified soybean cultivars and wild relatives to the aphid showed resistance to FA. The inheritance of the resistance gene and mechanism of action is not fully understood due to its complexity. Commercial production of soybean varieties with inherent resistance to FA has not been carried out due to the possible emergence of virulent FA biotypes that break down the dominant R gene. Therefore, pyramiding of multiple resistance genes in soybeans will help to increase the chances of sustainable production of soybean resistant to FA. Besides, more studies are required to fully understand the mechanism of resistance in soybean cultivars against FA to achieve crop improvement.

This research was funded by the National Research Foundation of Korea (Project No. RS-2023-00246459).