Adult nutrition affects reproduction and flight performance of the invasive fall armyworm,Spodoptera frugiperda in China

HE Li-mei ,JlANG Shan ,CHEN Yu-chao ,Kris A.G.WYCKHUYS ,GE Shi-shuai ,HE Wei,GAO Xiwu,WU Kong-ming

1 State Key Laboratory for Biology of Plant Diseases and Insect Pests,Institute of Plant Protection,Chinese Academy of Agricultural Sciences,Beijing 100193,P.R.China

2 College of Plant Protection,China Agricultural University,Beijing 100193,P.R.China

3 College of Plant Protection,Shenyang Agricultural University,Shenyang 110866,P.R.China

4 State Key Laboratory of Ecological Pest Control for Fujian and Taiwan Crops,Institute of Applied Ecology,Fujian Agriculture and Forestry University,Fuzhou 350002,P.R.China

5 College of Plant Protection,Henan Agricultural University,Zhengzhou 450002,P.R.China

Abstract Floral resources,such as carbohydrate-rich nectar or pollen,can bolster fitness and raise reproductive output of adult lepidopterans.Here,we used laboratory experiments to assess how those plant-derived foods impact adult fecundity,reproductive physiology and flight performance of an invasive strain of the fall armyworm,Spodoptera frugiperda (FAW;Lepidoptera:Noctuidae) in China.More specifically,supplementary feeding on bee pollen and honey enhanced FAW flight duration,testis size,ovarian development,longevity and adult fecundity.FAW adults attained the longest pre-oviposition(10.8 days) and oviposition period (6.8 days) and longevity (19.2 days) on 5% acacia honey.Upon access to 2.5% acacia honey and 2.5‰ pine pollen,S.frugiperda attained the highest mating rate (79.7%),fecundity (644.9 eggs/female) and egg hatching rate (82.3%).Feeding on honey further delayed decay of male testes,while ovarian development was enhanced when female moths were allowed access to 2.5% honey and 2.5‰ pine pollen.Upon feeding on 5% honey solution,S.frugiperda engaged in flight over the longest duration (9.5 h),distance (29.9 km) and speed (3.1 km h–1).Honey had a comparatively greater effect on the above parameters than pollen.Our findings help decipher FAW invasion patterns and population dynamics,facilitate the development of nutritional attractants,and contribute to integrated pest management of this newly-invasive pest in eastern Asia.

Keywords:supplementary nutrition,fecundity,adult reproduction,flight parameter,invasion biology,nutritional physiology

1.lntroduction

Carbohydrate-rich food items such as (extra-) floral nectar or pollen provide energy for insect development,mating,oviposition and flight (Hofereret al.2000; Lavanderoet al.2006; Sunet al.2009; Balzan and Wäckers 2013).Supplementary feeding on these resources can promote adult reproductive development,especially sexual maturity of ovary,raise fecundity (Thorpe and Caudle 1938; Elliset al.2005; Winkleret al.2006) and prolong longevity (Irvinet al.2007; Leeet al.2008).Both quantity and quality of adult food items can affect different life history parameters and dictate flight performance of migratory noctuid moths such asMythimnaseparata(Walker) andHelicoverpaarmigera(Hübner) (Wu and Guo 1997; Wanget al.2006).Yet,given their species-specific nutritional demands (Thompson 1999)and the variable accessibility,quantity,energy content and palatability of food items (Vattalaet al.2006; Fiedler and Landis 2007),insects exhibit marked preferences for pollen or nectar from certain plant species (Wäckers 2003;Winkleret al.2009; Andradeet al.2018).Adults ofHeliothis virescens(Fabricius) are able to differentiate among the three sugar sucrose,fructose and glucose (Ramaswamy 1987),while other species exhibit clear electrophysiological and behavioral responses to certain sugars,amino acids,allelochemicals and plant extracts (Blaney and Simmonds 1990).Hence,it is important to investigate the preference of a given insect species for particular food items and their ensuing effects on development,reproduction and longevity.Gaining these insights for both agricultural pests and insect natural enemies can be particularly valuable to advance integrated pest management (IPM) (Winkleret al.2010;Tenaet al.2015).

The fall armyworm (FAW),Spodopterafrugiperda(Smith)(Lepidoptera:Noctuidae),is a migratory pest native to the Neotropics,with strong flight capabilities (Smith and Abbott 1797; Luginbill 1928).As a polyphagous pest,FAW larvae feed on ＞350 different host plants belonging to 76 plant families,including several globally-important agricultural crops (Montezanoet al.2018).Since 2016,FAW has invaded and rapidly spread across large parts of Africa and Asia,inflicting important losses on local maize production(Goergenet al.2016; CABI 2017; Earlyet al.2018; Nakweta 2018; Sunet al.2021; Yanget al.2019).Considering howS.frugiperdahas historically been a key pest in the Americas,its morphology,host plant preference,migration and field colonization behavior,population genetics and mitigation options have been well studied (Yu and Mccord 2007; Barroset al.2010; Dumaset al.2015; Westbrooket al.2016; Midegaet al.2018; Guoet al.2019; Wuet al.2019; Zhanget al.2019).However,much of its nutritional ecology waits to be elucidated,especially for its invasive populations in eastern Asia.Earlier work on an invasive strain ofS.frugiperdain China has shown how larval diets impact adult fecundity and longevity (Heet al.2021).ThoughS.frugiperdaadults readily use carbohydrates for flight without converting them to fat (Van Handel and Nayar 1972),it remains unclear to what extent FAW adult feeding on particular plant-derived foods affects its reproduction and flight performance.

While honey is primarily composed of sugar (80%) and water (18%) (Solaymanet al.2016),pollen also contains proteins,amino acids and lipids and is highly valued by several insect species (Lundgren 2009; Areset al.2018).Lepidoptera adults feed primarily on sugar sources,and pollen-feeding behavior has only been described for few species (Gilbert 1972).Yet,lepidopteran adults do obtain vitamins and amino acids from (pollen-contaminated)nectar (Wäckerset al.2007).Noctuid moths visit flowers of a broad range of plant species including sunflowers,rapeseed,corn,pine trees and various members of the Rosaceae during migration (Liuet al.2016,2017; Changet al.2018).The flowers of these plants constitute important nectar sources that foraging noctuids can easily encounter and access in nature.However,to what extent foragingS.frugiperdaexploit these food items and derive fitness benefits from their consumption remains to be determined.

In this study,we systematically assess life history traits and reproductive physiology ofS.frugiperdaadults when fed with different types,concentrations and mixtures of honey or pollen under laboratory conditions.More specifically,we record dietary effects on mating,fecundity,egg hatching,longevity,flight capability and the development of (male,female) reproductive apparatus.Our findings aid the development of FAW nutritional attractants and contribute to the design of comprehensive IPM packages against this newly-invasive pest.

2.Materials and methods

2.1.lnsects

Laboratory trials were carried out during June–November 2019 at the Xinxiang Experiment Station of the Chinese Academy of Agricultural Sciences (CAAS; 35°18´13.71´´N,113°55´15.05´´E) in Henan Province,China.The strain used in the experiment is the same as that of Heet al.(2021).FAW individuals were field-collected in early 2019 in Dehong Autonomous Prefecture (Yunnan) and reared for more than five consecutive generations in the laboratory.Insects were kept at (25±1)°C,(75±5)% relative humudity (RH),and 16 h L:8 h D photoperiod.Larvae were fed on a soybean flour and wheat bran-based artificial diet (Greeneet al.1976),and 1st to 6th instar FAW larvae were reared in 22 cm×15 cm×8 cm plastic boxes.Once larvae were fully developed,they were transferred to plastic trays with vermiculite for pupation.On the 5th day after pupation,the female and male pupae were separated based on the method of Zhaoet al.(2011).

2.2.Adult fecundity

2.3.Observation of internal reproductive system

Testicular and ovarian anatomy were used to evaluate the effect of different food items on the internal reproductive system,for either unmated male or female FAW adults.The effect of food items on female ovarian development was determined forS.frugiperdaadults of varying age(i.e.,1-to 18-days old).Ovarian development (levels 1–5)was classified based on the standards of Zhang (2008)and Zhaoet al.(2019).We characterized the effect of the various food items on size (i.e.,major axis length) of male testes for adults of varying age (i.e.,1-to 15-days old) (Chenet al.2017; Heet al.2019).Moths were reared on nine food items (i.e., acacia honey,water,rape pollen,corn pollen,sunflower pollen,rose pollen,pine pollen,acacia honey+rape pollen and acacia honey+pine pollen)in 50 cm×50 cm×50 cm cages (200-mesh nylon,100 moths per cage) and kept at (25±1)°C and (75±5)% RH.Female moths were placed in a refrigerator at–20°C for 4 h before ovarian dissection,and male moths were immersed in 75%ethanol for 30 s before testis dissection.All moths were dissected under a stereomicroscope (TS-63X,Shanghai Shangguang New Optical Technology Co.,Ltd.,Shanghai,China) and the major axis length of testis was determined with OLD-SGD Show Software (Shanghai Shangguang New Optical Technology Co.,Ltd.,Shanghai,China).Each dietary treatment consisted of 30S.frugiperdamale or female moths (of a given age).

2.4.Flight performance

To recordS.frugiperdaadult flight parameters (i.e.,mean flight velocity,flight distance and duration),we employed a 24-channel,computer-monitored flight mill system (FXMD-24-USB type,Jiaduo Science Industry and Trade Co.,Ltd.,China) (Fuet al.2017).Each individual moth was weighed with an electronic balance (Mettler Toledo ME204; Beijing Haitian Youcheng Technology Co.,Ltd.,China) before and after the tethered flight assay.Individual moths were anesthetized with ether for 10–15 s and then attached by the tergal side of the mesothorax to the end of the flight mill arm using a 30-cm copper wire (0.3 cm in diameter).Moths were adhered to this wire using a drop of cyanoacrylate Super Glue (Deli Group Co.,Ltd.,Ninghai County,Zhejiang Province,China).Once a given moth was suspended from the flight mill arm,its wingbeat frequency was measured three consecutive times by using a stroboscope (Phaserstrobe PBX stroboscope; Monado Monarch,Amherst,New Hampshire,USA).The flight mill system was placed within a specially-designed climate chamber (MGC-450HP,Shanghai Yiheng Scientific Instrument Co.,Ltd.,China).Flight performance of individual moths was tested following their exposure to either of the nine food items (i.e.,acacia honey,water,rape pollen,corn pollen,sunflower pollen,rose pollen,pine pollen,acacia honey+rape pollen and acacia honey+pine pollen).FAW adults were exposed to a given dietary regime in 35 cm×35 cm×35 cm cages (200-mesh nylon,20 moths per cage) at (25±1)°C and (75±5)% RH.Next,3-day-old unmated male and female FAW adults were subject to tethered flight at 25°C and 75% RH,with each flight test starting at 22:00 for 12 h under continuous darkness.

2.5.Statistical analysis

The effect of dietary treatment onS.frugiperdaadult fecundity and flight performance was assessed using oneway analysis of variance (ANOVA),followed by a Turkey’s HSD (honestly significant difference) post-hoc test.Effects of dietary treatment and sex on FAW survivorship were appraised using Cox proportional hazards model (Wyckhuyset al.2008).Effects of diet and sex on flight performance,diet and age on male testis size or female ovarian development index were assessed using a two-way ANOVA followed by Turkey’s HSD post-hoc test.The differences of flight performance between female and male were analyzed by Student’st-test.Furthermore,to relate FAW flight duration and distance,Pearson linear correlation was used.In order to meet assumptions of normality and heteroscedasticity,proportional data were arcsine square root transformed.All data were analyzed using SPSS Statistical Software (IBM SPSS Statistics 20).

3.Results

3.1.Adult survival and reproduction

Dietary treatment affectedS.frugiperdadevelopment and longevity to variable extent (Table 1).Food items had a significant effect onS.frugiperdalifespan (χ2=8.080,df=1,P=0.004),sex (χ2=12.719,df=1,P＜0.001) and the sex×diet interaction (χ2=6.005,df=1,P=0.014) (Fig.1).Females attained the longest longevity on acacia honey,followed by acacia honey+rape pollen and sunflower pollen,and the shortest lifespan was recorded on rape pollen.Males also attained the longest lifespan on acacia honey,followed by acacia honey+rape pollen,and the shortest longevity was observed on rape pollen (Table 1).Food items also affected pre-oviposition (F8,411=5.383,P＜0.001),oviposition period (F8,411=6.545,P＜0.001),percentage egg-laying females (F8,18=13.707,P＜0.001),mating rate (F8,18=10.833,P＜0.001),fecundity (i.e.,eggs deposited per female,F8,411=4.311,P＜0.001) and egg hatching rate (F8,411=4.669,P＜0.001) (Table 1).On acacia honey,FAW adult attained the longest preoviposition and oviposition period,and the highest longevity,though egg hatching rate was the lowest.The highest female fecundity was attained for acacia honey+pine pollen-fed adults.While the lowest eggs deposited per female,oviposition period and mating rate were recorded in water-fed adults.The percentage of egg-laying females on diets containing honey(i.e.,acacia honey+pine pollen,acacia honey+rape pollen and acacia honey) was considerably higher than other diets,and the mating rate on diets containing honey and pollen (i.e.,acacia honey+pine pollen and acacia honey+rape pollen) was significantly higher than other dietary treatments.Diet did not affect the number of mating events.

3.2.Reproductive physiology

Female ovarian development was significantly affected by dietary treatment(F8,4697=70.416,P＜0.001),age (F17,4697=554.739,P＜0.001) and the diet×age interaction term (F136,4697=11.2000,P＜0.001) (Fig.2-A).Female ovarian development level increased significantly with the increasing age(Fig.2-C).FAW females obtained the highest ovarian development on acacia honey+pine pollen,followed by rose pollen,and acacia honey+rape pollen,while the lowest ovarian development was recorded on rape pollen,followed by water,then by pine pollen (Fig.2-E).Male testis size also was significantly affected by dietary treatment (F8,3768=23.231,P＜0.001),age (F14,3768=1 295.766,P＜0.001) and diet×age interaction(F112,3768=6.867,P＜0.001) (Fig.2-B).Male testis size decreased significantly with increasing age.Compared with 1-day-old males,testis size of 15-day-old males was 46.81% lower (Fig.2-D).FAW males attained the longest testis length on acacia honey+pine pollen,followed by acacia honey,and acacia honey+rape pollen,while the shortest testis length was obtained on pine pollen.In diets containing honey (i.e.,acacia honey+pine pollen,acacia honey and acacia honey+rape pollen),testis length was 1.96–5.42% longer than in those without honey (Fig.2-F).

Fig.1 Age-specific survival rate of female (A) and male (B) Spodoptera frugiperda adults when exposed to different food items.

3.3.Flight performance

Diet had a significant effect on weight loss during flight (F8,349=7.752,P＜0.001),as did sex (F1,349=3.923,P=0.048) and the diet×sex interaction term (F8,349=2.024,P=0.043)(Table 2; Fig.3-A).The lowest weight loss during adult(male and female) flight was recorded on acacia honey.Wingbeat frequency was not affected by diet alone,though significant effects were recorded for sex (F1,344=10.566,P=0.001),and diet×sex interaction (F8,344=4.281,P＜0.001)(Table 2; Fig.3-B).Females attained the highest wingbeat frequency on acacia honey,followed by acacia honey+rape pollen and corn pollen.Males on the other hand obtained the highest wingbeat frequency on pine pollen,followed by rape pollen and acacia honey+rape pollen.Diet (F8,351=12.568,P＜0.001) and sex (F1,351=7.037,P=0.008) had a significant effect on flight duration,however,no effects were recorded for diet×sex interaction (Table 2; Fig.3).Females attained the longest flight duration on acacia honey,followed by acacia honey+pine pollen and pine pollen,and the shortest flight duration was recorded on water.For male individuals,the longest flight was also registered on acacia honey,and the shortest flight on water.For flight distance,a significant effect was recorded for diet (F8,351=8.660,P＜0.001),though sex and the diet×sex interaction were not significant (Table 2; Fig.3).On acacia honey,the flight distance of females was significantly longer than that on other diets,and the shortest flight distance was recorded on water.For males,the longest and shortest flight distances were also recorded on acacia honey and water,respectively.Diet had a significant effect on flight velocity during flight (F8,351=2.752,P=0.006),as did sex (F1,351=0.194,P=0.660) and diet×sex interaction (F8,351=1.550,P=0.139) (Table 2;Fig.3).Females exhibited the fastest flight on acacia honey,while the highest flight velocity for males was recorded on sunflower pollen.Flight distance was positively correlated with flight duration (Pearson’s;r=0.942,P＜0.01).

4.Discussion

Adult consumption of carbohydrate-rich food items can bolster fitness,movement capabilities and reproductive output of numerous insect species (Lavanderoet al.2006;Lundgrenet al.2009; Balzan and Wäckers 2013),with variable,species-specific effects of certain food items.Especially for species with a long-lived adult stage,an extended oviposition period or (long-range) migration behavior,a sufficient and timely access to high-quality food resources is essential (Baoet al.2010).Here,we use laboratory assays to demonstrate how bee pollen and various types of honey enhance development and reproduction of the newly-invasive FAWS.frugiperda.Tethered flight trials further confirm how sugar-rich food items such as 5% acacia honey extendS.frugiperdamovement capabilities,and thereby amplify the dispersal range of this noctuid pest.

Fig.2 Effect of age and supplementary nutrition on female ovarian development (A,C and E) and male testis size (B,D and F)of Spodoptera frugiperda.Each bar reflects mean±SE and different lowercase letters reflect significant difference at the 5% level by two-way ANOVA followed by Turkey’s HSD.Sample sizes of index of ovarian development are 270 for each age and 540 for each food item,expect for 18-day-old and acacia honey+pine pollen (n=269 and 539,respectively); sample sizes of major axis length of testis are 262,261,261,265,256,263,259,256,264,248,256,255,257,269 and 270 from 1-to 15-day-old,and 435,434,420,422,444,429,432,445 and 442 from water to acacia honey+pine pollen.

Though lepidopteran species accumulate plant-derived nutrients during the larval stage,adult feeding is crucial to meet further resource needs for energy-demanding mating,oviposition,flight and other processes (Wu and Guo 1997;Jianget al.1999; Hou and Sheng 2000).Our work reveals how,in the absence of energy-rich foods,laboratoryrearedS.frugiperdaadults are still able to complete mating,oviposition and other reproductive behaviors.Yet,supplementary feeding on sugar-rich honey or pollen enhanced multiple reproductive parameters (e.g.,fecundity,mating frequency,oviposition period) and increased FAW lifespan.On 5% acacia honey,the oviposition period and lifespan of FAW adults increased with 2.61 and 5.22 days,respectively,as compared to distilled water.Moreparticularly,fecundity of FAW females fed on 2.5% acacia honey and 2.5‰ pine pollen solution increased 61.08%as compared to water-fed individuals.Similar to Fanget al.(2020),consumption of honey or sucrose solution thus prolongs FAW lifespan and increases oviposition.Similar findings were made for other (migratory) noctuids of agricultural importance,such asSpodopteraexempta(Walker),Spodopteraexigua(Hübner) andAthetislepigone(Moschler) (Gunn and Gatehouse 1987; Jianget al.2000,2015),underlining how carbohydrate-rich food items contribute to population build-up,foraging and migration processes -ultimately enhancing a species’ adaptability to variable or unpredictable environments.

Table 2 Flight performance of Spodoptera frugiperda adults (male and female) exposed to different dietary regimes,as assessed under laboratory conditions

An insect’s reproductive organs (i.e.,male testes,female ovaries) are critically important for mating,reproduction and population development.Ovarian dynamics can shed light upon the migration history and colonization trajectories of certain insect species (Jianget al.2011; Qiet al.2011).The development of the internal reproductive system of Lepidoptera can be divided into three types (Wanget al.2004),and our ovary dissections show howS.frugiperdaexhibits continuous egg development that is further enhanced through supplementary adult nutrition.Given that FAW ovaries remain poorly-developed on rape pollen,the type of pollen (and the carbohydrate content of food items)are likely important in shaping its reproductive development.Regarding the male reproductive organs,noctuids possess two symmetrical seminal nests at the larval stage that gradually fuse into a nearly spherical spermary (pre-pupal or early pupal stage).In adult noctuids,testis size decreases at variable pace with age (Milonas and Andow 2010; Chenet al.2017; Heet al.2019),and is further affected by adult nutrition,mating frequency,temperature and other biotic factors (Yeet al.2000; Milonas and Andow2010; Klemmeet al.2014).Our dissections indicate how adult nutrition equally shapes development of the male reproductive system inS.frugiperda.More specifically,honey-containing diets resulted in large-size testes of FAW males,with sugarrich food items possibly slowing testis decay (Milonas and Andow 2010; Chenet al.2017; Heet al.2019).

Aside from improving reproductive performance,adult nutrition also enhances movement capabilities,e.g.,flight performance of various insects (Wu and Guo 1997; Jianget al.2000; Geet al.2019).Our work confirms that certain food resources (especially honey) raise FAW adult flight velocity,duration and distance while decreasing flightinduced weight loss.Honey and pollen thus likely provide a blend of carbohydrates,proteins and lipids that help sustainS.frugiperdabiological activities with FAW exhibiting superior flight performance on 5% honey solution.For poorly dispersing lepidopterans,these kinds of insights potentially can be used to guide habitat management within ecologically-centered IPM programs (Gurret al.2017;Perovićet al.2018).As such,agro-ecosystems can be‘engineered’ and particular (flowering) plant species can be deployed within or near the agricultural field,to selectively discourage pest lepidopterans and disproportionately favor natural enemies (Lavanderoet al.2006; Villaet al.2017).Yet,considering how FAW is a long-range migrant with considerable dispersal capabilities (Nagoshiet al.2012),it is unclear what options might be available to individual farmers or land-managers to strategically deploy nectar-or pollen-bearing plant species,except for those with toxic or repellent nectar (Adler 2000).

Fig.3 Flight performance of Spodoptera frugiperda adults exposed to different dietary regimes.Different panels indicate adult weight loss during flight (A),wingbeat frequency (B),flight duration (C),flight distance (D) and flight velocity (E) of fall armyworm moths exposed to nine diets.Date are mean±SE.Female sample sizes of weight loss,wingbeat frequency,flight duration,distance and velocity are 31,27,17,16,21,22,19,17 and 19 from water to corn pollen,except for wingbeat frequency in sunflower and rape pollen (n=19 and 18,respectively);male sample sizes of weight loss, wingbeat frequency,flight duration,distance and velocity are 29,29,17,14,17,20,19,19 and 16 from water to corn pollen,except for wingbeat frequency in rose and sunflower pollen (n=16 and 18,respectively).Stastistically significant differences between male and female moths for a particular flight parameter are indicated with an asterisk (P＜0.05 level,Student’s t-test),while ns reflects nonsignificant differences.

FAW adults attain high fitness following consumption of various types of honey,pollen and honey-pollen blends.By integrating these findings with data on FAW behavioral orientation towards pheromones,light spectra,plantbased attractants or other food lures (e.g.,Tumlinsonet al.1986; Batista-Pereiraet al.2006; Carrollet al.2006),one can possibly devise ‘attract-and-kill’ or ‘attract-and-infect’system (Dembilioet al.2018; Gregget al.2018).These strategies can help lower the environmental footprint of pest management,broaden the action range of microbial pesticides and mitigate insecticide-resistant pest populations(Poullotet al.2001; Brandlet al.2017).Baits with pure hexose sugar,palm sugar,golden cane syrup or fermented molasses are attractive to noctuid moths -with acetic acid likely being a key attractive compound in fermentation products (Utrio and Eriksson 1977; EI-Sayedet al.2005).Mixtures of acetic acid and 3-methyl-1-butanol further attract various lepidopteran species such asMamestra configurata(Walker),Xestiac-nigrum(L.),Mythimna unipuncta(Haw.),Hypenahumuli(Harris),Lacanobia subjuncta(Grote & Robinson),Pyralisfarinalis(L.) orAnagastakuhniella(Zeller) (Landolt 2000; Landoltet al.2007; Tóthet al.2010).By integrating our data with some of the above findings,traps can be designed to monitor,catch,infect and eventually killS.frugiperdaacross its newly-invaded range in eastern Asia.

5.Conclusion

FAW,S.frugiperda,has fast become an agricultural pest of global allure,threatening the sustained and profitable production of corn,rice,wheat and other crops in the Americas,Africa and Asia.Our work illuminates how floralderived diets impact development,fecundity,longevity and flight capability ofS.frugiperdaadults.In particular,the addition of honey to adult diets helped delay testis decay,prolonged the oviposition period and lifespan,raised fecundity and boosted flight performance.Our work contributes to an improved understanding ofS.frugiperdanutritional ecology,provides baseline information to design nutritional attractants and helps finetune IPM approaches for this pest in its invasive range in eastern Asia.

Acknowledgements

This work was supported by the earmarked fund of China Agriculture Research System (CARS-15-19) and the Central Public-interest Scientific Institution Basal Research Fund,China (Y2019YJ06).

Declaration of competing interest

The authors declare that they have no conflict of interest.