Asian Citrus Psyllid: Biology, Ecology and Management of the Huanglongbing Vector

By SANDRA A ALLAN, SIDNEY ALTMAN, ANTONIO JULIANO AYRES and 26 more

Asian citrus psyllid (ACP), Diaphorina citri, is an insect pest which transmits a bacterium, Candidatus Liberibacter asiaticus (CLas), primarily through feeding in newly emergent foliage of citrus trees. This pathogen causes a disease known as Huanglongbing (HLB), or citrus greening, which has become the most debilitating and intractable disease in citrus crops.

This book, written by a team of experts on the Asian citrus psyllid, gathers together everything currently known about the biology and ecology of this important pest species, examines the transmission and acquisition processes of the pathogen, and looks at current management practices and their effectiveness. The potential for new, innovative management techniques are also described, along with the economic implications of managing this rapidly establishing disease.

This book:

Covers all aspects of Asian citrus psyllid biology and ecology for the first time in one place.
Examines new, innovative management practices and assesses their effectiveness.
Discusses the vector-pathogen relationship in detail.
Explains the economics of controlling this devastating pest.

This title is essential reading for all researchers involved in the management and control of Asian citrus psyllid, extension agents, and pest management consultants. It will also be of great use to graduate students in applied entomology and related disciplines.

• Language: English
• Publisher: CAB International
• Release date: Jun 11, 2020
• ISBN: 9781786394101

Book preview

1 Asian Citrus Psyllid Life Cycle and Developmental Biology

David G. Hall

*

US Department of Agriculture – Agricultural Research Service, Fort Pierce, Florida, USA (Retired)

* Email: [e-mail]

Considerable research and review information has been published on the biology of the Asian citrus psyllid (ACP), Diaphorina citri. Hussain and Nath (1927) laid the foundation for present knowledge of ACP biology in an early comprehensive review upon which many advances have been made. Presented here are highlights of the ACP life cycle and developmental biology. ACP favors tropical/subtropical climates and hot, coastal zones (Catling, 1970; Hodkinson, 2009; Jenkins et al., 2015). Typical of all Hemiptera, ACP undergoes simple (incomplete) metamorphosis with the three typical life stages: egg, nymph and adult. Citrus is regarded as a primary host plant of the psyllid and the most important from an economic standpoint, but a number of other species within the plant family Rutaceae, subfamily Aurantioideae, are utilized by the psyllid for food and reproduction. The psyllid’s reproductive biology is closely synchronized with the production of shoots of new leaf growth (flush), as oviposition occurs exclusively on emergent leaves (often called feather flush), sometimes including young leaves associated with emergent floral shoots (Hall et al., 2008a), and young, unhardened leaves are required for the development of nymphs. While immatures of some species within the Psylloidea, including the African citrus psyllid Trioza erytreae, develop in pit-like deformations or galls induced on leaves, the Asian citrus psyllid does not and thus is free-living throughout its development on a flush shoot. However, many eggs and early instar nymphs are usually protected or hidden from view within unexpanded leaves or clusters of young leaves.

1.1 Adult Reproductive Biology, Life Characteristics and Polymorphisms

The ACP is a bisexual species, with equal numbers of females and males observed in some populations (Aubert and Quilici, 1988; Tsai and Liu, 2000; Nava et al., 2007) and a predominance of females in others (Pande, 1971; Hodkinson, 1974; Alves et al., 2014). Temporal emergence patterns of males and females are similar, with no evidence of protandry or protogyny (Wenninger and Hall, 2007; Hall and Hentz, 2016). Adults are small (2.7–3.3 mm long) with mottled brown wings (Fig. 1.1A). The end of the male’s abdomen bends upward while the end of the female’s abdomen is straight and pointed (Husain and Nath, 1927) (Fig. 1.1B). Adults rest or feed on plants with their bodies characteristically held at a ~45° angle (range 30–60°) to the plant surface. Adults feed on young stems and on leaves of all stages of development but preferentially move to newly developing flush to feed, mate and oviposit. Mating may primarily take place on flush shoots where females feed and lay eggs, but both sexes frequently walk on limbs and branches in the interior of a tree where they can sometimes be found mating. Adult males and females locate mates, in part, using substrate-borne vibrational sounds (Wenninger et al., 2009a). These sounds cannot be detected by the human ear, but individuals calling mates can be seen rapidly vibrating or beating their wings for short periods of time. Behavioral evidence indicated that females emit a sex pheromone (Wenninger et al., 2008), and recently Zanardi et al. (2018) identified acetic acid as possibly being involved. In addition, female-produced cuticular hydrocarbons may function as sex pheromones when males are in close proximity (Mann et al., 2013; Martini et al., 2014a; Moghbeli et al., 2014). During copulation, a male and female are positioned side by side with their heads facing the same direction, the male bending the tip of his abdomen down to the female. He uses his legs on the side next to the female to hold her while supporting himself on the plant surface with his legs on the other side (Hussain and Nath, 1927). Wenninger and Hall (2007) reported that copulation lasts from 20 to 100 min and occurs predominantly during daylight hours. Pande (1971) reported that mating takes place at any time during the day or night. Females maintain optimum reproductive output by mating multiple times with the same or different partners (Wenninger and Hall, 2008a).

Fig. 1.1. (A) An adult Asian citrus psyllid, Diaphorina citri. (B) Backlight illumination can be used to distinguish females from males based on the tip of their abdomens. The upper adult is a female, the lower adult is a male. (C) Asian citrus psyllid eggs. (D) The five nymphal instars of the Asian citrus psyllid.

Adults exhibit three relatively distinct abdominal colors: gray/brown, blue/green and orange/yellow (Husain and Nath, 1927; Wenninger and Hall, 2008b). Most individuals within Florida populations are blue/green, while gray/brown individuals are rarest (Hall and Hentz, 2016). Age-related shifts may occur over time in an individual’s color, but are not seasonal. The biological significance of these polymorphisms is slowly being unraveled. Husain and Nath (1927) reported that the abdomen of gravid females turns distinctively orange, particularly during spring. Wenninger and Hall (2008b) reported that abdominal color has little value as an indicator of sexual maturity and only limited value for discerning female mating status. The orange/yellow color in females reflects the presence of eggs in the abdomen; in males, it seems to derive from the color of the internal reproductive organs and this color is generally only expressed in older males. Females may associate male color with reproductive success, as they avoid blue males after previous experience (Stockton et al., 2017). Orange males mate more frequently than blue males and appear to be more sexually aggressive in mating attempts (Stockton et al., 2017). Interestingly, females that mated with orange males laid twice as many eggs as those mated to blue males (Stockton et al., 2017). There is evidence that blue/green individuals are more apt for long-distance dispersal (Martini et al., 2014b), which could be related to their larger size (Paris et al., 2016). Differences have been reported among color morphs with respect to insecticide resistance (Boina and Bloomquist, 2015). Hemocyanin may in part be responsible for the blue/green morph (Ramsey et al., 2017), but it is not known why some psyllids might produce more hemocyanin than others.

Husain and Nath (1927) reported that new adults in the Punjab region of what is now Pakistan began copulating soon after emergence and females began laying eggs on citrus soon afterwards following a pre-oviposition period of 1–3 days. In nearby Rajasthan (India), Pande (1971) reported that adults copulated 12–60 h after emergence and that oviposition commenced 8–20 h later, indicating a pre-oviposition period of 0.8–3.3 days. Wenninger and Hall (2007) reported that newly emerged adults in Florida at 26°C on orange jasmine (Murraya paniculata) mated within 2–3 days with oviposition beginning 1 day after mating for a pre-oviposition period of 3–4 days. Contrasting observations in Brazil by Alves et al. (2014) and Nava et al. (2007) indicated a pre-oviposition period of 8.5–10.9 days at 24–25°C on orange jasmine. Clearly, there are some discrepancies among pre-oviposition periods reported for ACP that may be due to environmental and host plant factors. Based on data from Yang (1989), changes in photoperiod and light intensity can influence the pre-oviposition period. Uechi and Iwanami (2012) noted that maturation of a female’s ovaries was faster when new adults fed on younger leaves.

Females lay eggs throughout their lives, provided that tender flush is available. Adult females typically lay 500–800 or more eggs over their lifetime (Husain and Nath, 1927; Tsai and Liu, 2000; Nava et al., 2007), with a reported maximum of 1378 (Tsai and Liu, 2000; a maximum of 1900 referenced by Tsai and Liu was apparently an error). The number of eggs laid by an individual female may vary depending on factors such as temperature and host plant species (Liu and Tsai, 2000; Tsai and Liu, 2000; Nava et al., 2007; Westbrook et al., 2011; Alves et al., 2014; Hall and Hentz, 2016; Hall et al., 2017a). Adult females held at 25°C were reported to lay an average of 858 eggs on grapefruit (Citrus paradisi) compared with 572, 613 and 626 on rough lemon (Citrus jambhiri), sour orange (Citrus aurantium) and orange jasmine, respectively (Tsai and Liu, 2000). Differences in oviposition rates reported on different host plant species (e.g. Tsai and Liu, 2000; Nava et al., 2007; Westbrook et al., 2011; Hall et al., 2017a) may be attributed to differences in plant volatiles, secondary plant compounds, nutritional quality and other factors but not in the abundance of simple foliar trichomes (Hall et al., 2017b). At 25°C, oviposition rates per female steadily increased over the first 10 days after mating, reaching a peak within 12–18 days, after which rates steadily declined on orange jasmine and grapefruit. In contrast, oviposition rates on lemon and sour orange peaked at about the same time but then remained somewhat steady through 60 or more days (Tsai and Liu, 2000). Nava et al. (2007) reported that females laid the majority of their eggs during the first 10 days after oviposition commenced. There can be substantial day-to-day variation among individual females in numbers of eggs laid (Husain and Nath, 1927). Such variation can be a result of mate availability, male color morph, age and possibly other biotic or abiotic factors. Based on Nava et al. (2007), differences in fecundity may also have a genetic basis.

A number of factors may influence longevity of adult ACP, including temperature, relative humidity, host plant species, food availability and reproductive status (Tsai and Liu, 2000; McFarland and Hoy, 2001; Nava et al., 2007; Hall et al., 2008b). Husain and Nath (1927) reported that adults live as long as 2 months or more, during which females may continually lay eggs. These authors reported that one adult lived for 189 days. Pande (1971) indicated that female ACP generally live longer (mean 45.0 days) than males (mean 40.5 days). Nava et al. (2007) reported that at 24°C adult males lived an average of 21–25 days and females lived an average of 31–32 days on lime (Citrus limonia), Sunki mandarin (Citrus sunki) and orange jasmine. Longevity was one of several factors that collectively suggested a greater importance of females compared with males in the epidemiology of huanglongbing (Hall, 2018). Females held at 25°C lived an average of 40–48 days on rough lemon, sour orange, grapefruit and orange jasmine (Tsai and Liu, 2000). Adult psyllids can survive without food for at least several days, depending on environmental conditions (Hall and McCollum, 2011). They may sustain themselves for 3–8 days or more on some plant species outside of the Rutaceae such as cotton (Gossypium hirsutum), guava (Psidium guajava) or tomato (Solanum lycopersicum) (Hall et al., 2008b).

1.2 Development of Eggs and Nymphs

Eggs are oval, clear to light yellow when freshly deposited and bright yellow-orange with two distinct red eye spots at maturity (Fig. 1.1C). Eggs are laid on terminal growth of newly developing plant tissue, including leaf folds, petioles, axillary buds, upper and lower surfaces of young leaves and tender stems (Tsai and Liu, 2000). The average size of an egg was reported by Tsai and Liu (2000) to be 0.31 mm long and 0.14 mm in diameter. Eggs are anchored to young flush shoots with a tapered basal stalk (pedicel) at the posterior/lower end averaging 0.038 mm in length (Husain and Nath, 1927). Once embedded in plant tissue, the pedicel facilitates water exchange with the plant, which is essential for egg development (Burckhardt, 1994). Means of 16–27 eggs per shoot have been reported along with a maximum of 777 eggs on one single flush shoot (Hall et al., 2008a). If young flush is not available, gravid females may discharge eggs, singly or in groups, on to the surface of a plant without imbedding the stalk. These eggs fail to develop.

The ACP develops through five nymphal instars (Fig. 1.1D). Early instars are largely sedentary and move only when disturbed or overcrowded (Tsai and Liu, 2000), whereas older nymphs are more mobile. Nymphs feed on young leaves and stems, continually secreting copious amounts of honeydew in conjunction with a white wax-like material (Tsai and Liu, 2000; Ammar et al., 2013). Adult females also produce a similar white excretory substance, but males only produce clear sticky droplets (Hall et al., 2010). Honeydew associated with large infestations of nymphs collects below infested flush on which black sooty mold can develop.

Husain and Nath (1927) and Tsai and Liu (2000) presented information on distinguishing the five instars. The first-instar nymph is 0.3 mm in length and 0.17 mm in width with a light pink body and red compound eyes. The second instar is 0.45 mm in length and 0.25 mm in width; rudimentary wing pads are visible on the dorsum of the second instar’s thorax. The third instar averages 0.74 mm long and 0.43 mm wide, with well-developed wing pads and evidence of antennal segmentation. Third-instar nymphs have a single seta on each antenna. Fourth instars average 1.01 mm in length and 0.7 mm in width, with mesothoracic wing pads extended to the compound eyes and metathoracic wing pads extended to the third abdominal segment. The fourth instar has two setae on each antenna. Fifth instars average 1.6 mm long and 1.02 mm wide, with the mesothoracic wing pads extended toward the front of the compound eyes and the metathoracic wing pads reaching the fourth abdominal segment. Three setae are found on each antenna of fifth-instar nymphs. In some mature nymphs, the abdominal color turned bluish green while in others the abdomen turned pale orange (Tsai and Liu, 2000), but the significance of these color changes is not known.

The ACP is a multivoltine species and so the number of generations produced each year varies depending on temperature and other environmental factors in conjunction with the availability of young flush for oviposition. Husain and Nath (1927) and Pande (1971) reported nine to ten generations annually, with as many as 16 generations observed by Atwal et al. (1968). Young trees may continually produce at least some new growth over much of the growing season, supporting many generations each year. Older trees usually flush at certain times of the year and, after a flush, adults may be stimulated to disperse from these trees in search of new growth to support reproduction. Population levels of the ACP in Florida are usually highest during late spring or early summer before the rainy season and lowest during winter. However, large populations of eggs, nymphs and adults can occur at any time of year, depending on environmental conditions and the presence of young flush (Tsai et al., 2002; Hall et al., 2008a). Diapause has not been reported in ACP (Burckhardt, 1994).

1.3 Temperature Effects

The effects of temperature on ACP longevity, reproduction and development have been studied in detail under laboratory conditions (Liu and Tsai, 2000; Mizuno et al., 2004; Fung and Chen, 2006; Nakata, 2006; Nava et al., 2007; Hall et al., 2011; Hall and Hentz, 2014). Table 1.1 presents a summary of published information pertinent to the biology of the psyllid reared on orange jasmine at temperatures of 15–33°C. Nava et al. (2007) reported female longevity on ‘Rangpur’ lime to range from a mean of 88.3 days at 15°C to 28.7 days at 33°C. Liu and Tsai (2000) reported that maximum female longevity ranged from 117 days at 15°C to 51 days at 30°C. Data from Wenninger and Hall (2007) suggested mating activities paused in a greenhouse during mid-afternoon when air temperatures reached 40°C. Lower and upper temperature thresholds for oviposition were reported as 16.0°C and 41.6°C, respectively, with 29.6°C estimated as optimal (Hall et al., 2011). Skelley and Hoy (2004) reported a cessation of oviposition following an air-conditioning failure causing the temperature in a rearing room to remain at 34°C for 5 days. Oviposition gradually resumed over 2–3 weeks once temperatures went back to normal. Fecundity, survival from egg to adult and development time vary with temperature and among host plant species (Tsai and Liu, 2000; Alves et al., 2014). Lower and upper temperature thresholds for development were reported to be 10.9–11.7°C and < 33°C, respectively (Liu and Tsai, 2000). The optimal temperature range for ACP development on orange jasmine is generally around 24–28°C based on fecundity, survivorship and speed of development and closer to 28°C based on intrinsic rates of increase (Rm) (Table 1.1). However, only 75–84% of immatures survived to the adult stage within this optimum temperature range, even under fairly ideal laboratory conditions (Liu and Tsai, 2000). The majority of immatures failing to reach the adult stage died during the egg and first nymphal instar stages.

Table 1.1. Biology of Diaphorina citri on orange jasmine at different constant temperatures.

Linear regressions relating developmental rates of immatures to air temperature have facilitated estimates of developmental thresholds and degree-day growth models for the psyllid (Liu and Tsai, 2000; Fung and Chen, 2006; Nava et al., 2007; Milosavljević et al., 2018). Information has been published on life-table parameters for the psyllid, including intrinsic rates of increase, net reproductive rates, mean generation times and population doubling times at different temperatures and on different host plants (Tsai and Liu, 2000; Fung and Chen, 2006; Nava et al., 2007; Alves et al., 2014).

ACPs occur in some of the hottest climates in which citrus is grown and are known to survive at least short exposures to temperatures as high as 45°C in arid climates (Aubert, 1988, 1990). Mortality due to high temperatures may be intensified when it occurs in conjunction with low humidity producing a high saturation deficit index (SDI) (Hodkinson, 2009). Research on other psyllid species has shown that high SDI may result in mortality, reduced fecundity and slower rates of development. However, Hall and Hentz (2014) found that adult ACPs were less tolerant of high temperatures (45°C) when humidity was moderate (75%) than when it was low (23%). Heat acclimation helps explain why the psyllid can survive in some geographical areas where afternoon air temperatures sometimes exceed 40°C for several hours. Atwal et al. (1968) reported that 5th-instar nymphs and adults withstood 45°C for up to 4 h and adults withstood 40°C for up to 12 h. Heat shock proteins probably help psyllids survive high temperature extremes (Marutani-Hert et al., 2010). ACPs are common and abundant in Pakistan and yet daily air temperatures during the summer may exceed 40°C for 7–8 h, with peaks as high as 44°C with 33–39% relative humidity (RH) (Hall and Hentz, 2014). Levels of mortality or adverse biological effects from such temperature extremes are not known, but ACP populations remain prevalent in the Punjab region of Pakistan (Hoddle and Hoddle, 2013). Some eggs and nymphs of the ACP may escape high lethal air temperature events because they are protected within clusters of young flush leaves.

In subtropical areas with seasonal changes in climate, psyllids infesting citrus may sometimes be subjected to freezing temperatures for various periods of time. ACPs are known to survive at least short exposures to temperature extremes as low as –7°C in subtropical wet areas (Aubert, 1988, 1990). Atwal et al. (1968) reported that fifth-instar nymphs and adults exposed to 0°C died within 6–8 h. Working with psyllids reared under greenhouse conditions at ~25°C, 66–48% of adults survived exposure for 2–3 h to temperatures of –5 to –6°C; greater than 60% of nymphs survived 8–10 h of exposure to temperatures as low as –4°C; and around 50% or more eggs remained viable after being exposed for 2–3 h to temperatures as low as –8°C (Hall et al., 2011). The psyllid can acclimate to colder temperatures and subsequently survive some freeze events that would otherwise be lethal (Hall et al., 2011). According to Hodkinson (2009) and others, insects can avoid lethal freezes by lowering the ‘super cooling point’ of their body tissues. It is likely that some adult psyllids may survive a freeze by finding cold protection in bark crevices or within ground litter. Some freeze events might not be severe enough to kill a large percentage of psyllids but may indirectly cause mass mortality of eggs and nymphs by killing young flush. Young citrus flush shoots have been noted as being fairly tolerant of a 3 h freeze at –1.7°C or a 30 min freeze at –2.2°C, but large percentages of new shoots may die following freezes of –2.2°C for 2–3 h or following a –3.3°C freeze for 30 min (Oswalt, 2008).

1.4 Humidity, Rain and Sunlight

Most laboratory investigations on ACP biology have been conducted at constant relative humidity levels in the 60–80% range. In Florida, where high population levels of the psyllid are known to occur, humidity during the night and early hours after daybreak usually approaches 90% or higher. Skelley and Hoy (2004) reported on a rearing procedure for the psyllid in which RH in an air-conditioned rearing room varied seasonally from 35% to 65%, noting that females produced fewer eggs when the RH in the room dropped below 40% (information was not provided on humidity levels in the canopies of rearing plants). McFarland and Hoy (2001) reported that, in the absence of a host plant, adult longevity decreased as RH was incrementally decreased from 97% to 7% at either 25°C or 30°C. The effect was more pronounced at 30°C, at which about 80% adults survived for 20 h at 97% humidity, compared with 0% survival at 7% humidity. With respect to rain, Aubert (1987) speculated that monthly rainfall in excess of 15 cm was generally associated with low populations of the psyllid due to eggs and young nymphs being washed off plant surfaces. However, according to Husain and Nath (1927), because eggs are anchored into plant tissue by their stalks, they cannot be washed off by rain. Population levels of eggs, nymphs and adults did not appear to be negatively influenced by rain in two Florida orchards where rainfall sometimes exceeded 9 cm a month (Hall et al., 2008a).

Adult activity (flight, walking, feeding, mating and oviposition) is prompted by daylight. Flight activity varies with changes in sunlight (Hall, 2009) and is pronounced during warm, sunny afternoon hours (Aubert and Hua, 1990; Sétamou et al., 2012; Paris et al., 2015). Flight initiation by ACP is influenced by changes in barometric pressure and increases in air temperature, but changes in humidity do not affect its dispersal (Martini and Stelinski, 2017). The psyllid is positively phototaxic (Husain and Nath, 1927; Aubert and Hua, 1990) and its behavior is strongly influenced by sunlight (see Hall et al., 2008b, 2018). Paris et al. (2017) showed that positive phototaxis by walking psyllids was associated with short-wavelength ultraviolet (UV) light (350–405 nm), while little or no walking responses were observed at longer wavelengths in the visible spectrum from green to yellow to orange (500–620 nm). Increased UV light (high spectral irradiance in the 250–400 nm range) was one of several factors speculated as possibly being responsible for lower population levels of the psyllid at higher altitudes (Jenkins et al., 2015), but the effects of high UV on psyllid reproduction, development and longevity are not known.

Published research findings on ACP biology under laboratory conditions are usually accompanied by information on photoperiod, which is usually in the range of 13–16 h of daily illumination. However, irradiation and light intensity are generally not reported, although light type and position sometimes are. For example, Skelley and Hoy (2004) reared the psyllid in rooms illuminated only by fluorescent lamps (5000–6000 lux output) suspended 2.5 cm above rearing cages. Hall et al. (2016) illuminated ACP-infested plants in the laboratory using light-emitting diode lamps (PAR38 22º, 17 W, white 3000K floodlights) positioned 45–60 cm above the plants, and in a walk-in chamber using 400 W mercury vapor lamps positioned 23 cm above rearing cages. Wenninger et al. (2009b) reported working at a light intensity of about 3200 lux (provided by fluorescent lamps) recorded just above rearing vials. Good comparisons remain to be made of ACP biology under fluorescent lamps, mercury lamps, light-emitting diodes, halogen and tungsten lamps versus sunlight. Hall and Hentz (2016) considered cooler temperatures during winter as primarily responsible for slower development and reduced productivity of ACP reared on different host plant species, although shorter photoperiods and reduced solar radiation/light intensity may also have been factors. Data presented by Yang (1989) indicated that fecundity increased with light intensity (1200–15,000 lux) and photoperiod (6–18 h daily illumination), and that these parameters influenced the pre-oviposition period. Information on solar radiation as it relates to ACP reproduction potential and intrinsic rates of increase is needed. Mass-producing of ACP would benefit from expanded information on the influence of light on the reproductive biology of the psyllid. To this end, Hall and Hentz (2019) reported that photoperiod, irradiance and illuminance were positively correlated with ACP reproductive rates.

1.5 Expanding the Knowledge Base

Newly emerging research interests are continually expanding our knowledge of the biology of the ACP. Endosymbionts such as Wolbachia, Profftella and Carsonella may affect certain parameters of psyllid reproduction and biology (Hoffmann et al., 2014; Ramsey et al. 2017). Much remains to be discovered regarding the developmental biology of ACP infected by ‘Candidatus Liberibacter asiaticus’ (CLas), the bacterial pathogen responsible for huanglongbing. Evidence has already been presented that this pathogen increases ACP dispersal potential (Martini et al., 2015) and susceptibility to insecticides and entomopathogens (Tiwari et al., 2011; Orduño-Cruz et al., 2016). Ren et al. (2016) reported that the bacterium had obvious effects on the biology of the psyllid, including increased fecundity and longevity of females. Pelz-Stelinski et al. (2010) and Pelz-Stelinski and Killiny (2016) found that females carrying the pathogen were more fecund than uninfected counterparts and that developmental rates of infected nymphs were reduced, but that longevity of infected adults was reduced. Electropenetrography showed that ACP infected by the pathogen foraged more often than healthy ACP (Killiny et al., 2017). Transcriptome and proteome analyses on ACP infected by CLas showed an upregulation of transcripts and proteins involved in defense and immunity (Kruse et al., 2017; Ramsey et al., 2017). Further advances in our knowledge of ACP biology will be beneficial with respect to finding novel methods of managing vector populations and thus huanglongbing.

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2 Functional Anatomy of the Asian Citrus Psyllid

Joseph M. Cicero

*

Department of Entomology and Nematology, University of Florida, Gainesville, USA

* Email: [e-mail]

2.1 Introduction

Classical authors made huge and valuable baseline contributions to understanding homopteran anatomy for the 80 or so years before quantum improvements in technique (Karnovsky, 1965; Friedrich et al., 2014) and resolution of microscopy (Palucka, 2002) arrived. However, many aspects of their works, along with their crude sketches and interpretive skills, have been grandfathered into modern literature without validation using modern instruments, techniques or rigor. Further, a homological school of thought presided for use in determining identifications, functions and evolution of anatomical components. Understandably, this school persists today, even though advancements in cell biology with special reference to cytological and histological miniaturization, reduction and simplification (Beutel and Haas, 1998; O’Malley et al., 2016; Randolf et al., 2017) make it very difficult to maintain the school while at the same time ushering in a new era that caters to protein chemistry, molecular biology, endocrinology and pest control. In fact, review of the classical homopteran literature led to the suggestion that the difficulty with maintaining the school is because the task of correlating certain anatomies from one taxon to another is actually a phylocytological problem (Cicero and Brown, 2012). Nevertheless, on review of the ACP literature, it is clear that a modern working system of descriptive, structural terminology for homoptera is still necessary and in dire need. An example solution to this problem would be a homopteran anatomy ontology modeled from the successful Hymenoptera Anatomy Ontology Project (Yoder et al., 2010).

As of this writing, a search of prominent databases for articles using ‘Diaphorina’ and ‘citri’ as Boolean keywords yielded surprising results (Table 2.1). Excluding the taxonomic and pre-1970 literature, only ten monographic publications elucidating aspects of external anatomy (morphometrics, five) and seven elucidating aspects of internal anatomy were garnered from the total number of hits. The rest were mostly insect pest management, ecology, pathology and molecular. It is clear from these statistics that anatomy of ACP falls far short of critical need. In fact, most of what can be assembled regarding ACP functional anatomy is transferred, or inferred, from studies of other species, especially the potato psyllid, Bactericera cockerelli (Sulc) (PoP).

Table 2.1. Hits from the Boolean keyword search ‘Diaphorina’ AND ‘citri’ using three prominent US databases (last accessed: September, 2018).

As a summary chapter of ACP anatomy has already been published (Brown et al., 2016), and monographs of ACP anatomy are few, this chapter’s sections will summarize core knowledge of select anatomical components and, where possible, extend them into a prospectus of ideas that critically need consideration during the eventual movement to Gupta’s (1994) ‘molecular morphology’.

2.2 Embryology

No modern studies of ACP embryology (Fig. 2.1) are currently available. Therefore, the general aspects of psyllid embryonic development are only inferential from those determined by modern techniques for relatives such as aphids, Oncopeltus, Rhodnius and others.

Fig. 2.1. Post-katatreptic embryo of ACP. Prepared by phase partition fixation (Zalokar and Erk, 1977) using 8% aqueous formaldehyde and heptane. Abdominal segmentation is not well differentiated. Line = 50 μm.

Of special interest in ACP embryology is the biogenesis of the stylets (Section 2.3.5) and biogenesis of the filter chamber (Section 2.4.2). Embryonic biogenesis of the configuration of hypodermal cells needed to secrete and house presumptive (new) stylets of the true first instar (psyllids may have pre-eclosion instars with their own molting events) should have marked differences from that of consecutive pharate instars insofar as the first embryonic process probably does not involve an end-cap, and almost certainly does not involve an embedded matrix. Concordantly, the filter chamber is mostly made up of midgut tissue and, reasonably assuming that first instars (‘crawlers’) have one, then in theory embryogenesis of the midgut should, to a certain extent, follow the primitive motif of initiation between the stomodeal and proctodeal invaginations, then depart from that motif to allow co-location, convolution and ensheathing of its opposing ends to form one. This process would be in contrast to the direct inheritance of the filter chamber from one instar to the next. The departure is of great interest because it is the ontogenetic stage in which study of filter chamber biogenesis and evolution is best undertaken.

2.3 Oral Region

The oral region of PoP was defined as internal and external anatomy occurring between the anterior margin of the ‘rostrum’, a ventral, tear-shaped sclerite, and the profurcasternum, located at the base of the labium (Cicero et al., 2015) (Figs 2.1, 2.2). The ACP oral region is consistent with this PoP model. In both, the labium is shaft-like and evolutionarily posteriorized to where it is now directed ventrally between, or slightly posterior to, the procoxae, a condition considered to be derived for Sternorrhyncha (e.g. scales, larval whiteflies, psyllids) (Hennig, 1981:248; Grimaldi, 2003:340). This posteriorization coevolved with, and facilitated, the reflexing of oral region components primitively dorsal to it (maxillae, mandibles, labrum and clypeus) to their current position along the ventral face of the cranium so that the mandibular stylets (‘mandibulars’) are ventral to the maxillary stylets (‘maxillars’) (Fig. 2.3).

Fig. 2.2. Oral region of adult ACP. The rostrum is ventral to structures shown and fully out of view. Tentorial arms, or simply called ‘side arms’, are graphically drawn in place (*) since they were broken away to access the mandibular and show its inner lateral surface (Section 2.3.5). Profurcasternal sclerites were torn away during extirpation. The crumena redirects the stylet bundle from a posterior direction to a ventral direction for direct address into the host plant. The auricle is an inner lateral extension of the ‘flat’ or basal terminus (cf. Fig. 2.3). Line = 100 μm. Inset: magnification of mandibular showing that all but the distal end of the loading sleeve was torn away during extirpation. The cushion collects at a critically significant space inside the hollow, at the inner lateral surface of the proximal base of each stylet (encircled). This position is directly posterior to the auricle (cf. Fig. 2.5C and Section 2.3.5). Line = 10 μm.

Fig. 2.3. Adult ACP. Schematic showing the routing of stylets from their basal termini to exit from the crumena. The stylet ‘base’ is the length inside the head. The stylet ‘shaft’ is the length outside the head. Not drawn to scale – maxillar interlock and mandibular rerouting actually occur very near exit of the stylet bundle through the preoral orifice. The stylet basal terminus is the ‘flat’. Its auricle protrudes in an interior direction (cf. Fig. 2.2). Cross-section (xs) indicates two loci representing the intersected edges of the basal terminus or ‘flat’ of the stylet (cf. Fig. 2.5C). With the head reflexed, the mandibulars are ventral to the maxillars at their basal termini, but they are rerouted to a position lateral to the maxillars, then the stylet bundle as a whole is rotated 90°.

In the primitive condition, the outer lateral mandibular and maxillar surfaces were exposed to the outside. However, this reflexing or ‘ventralization’ (‘opisthognathy’ or ‘opisthorrhynchy’), included closure of the exposure by modification of external lateral head structures (e.g. the gena, lora, maxillary and mandibular plates of various authors) such that only a minute opening, the ‘preoral orifice’, remained (Fig. 2.3). It also included modification of the precursors of the stylets into very long, highly specialized feeding devices that pass through this tightly fitting orifice. In simplest terms, this explains how the stylet bases and the true mouth were deeply internalized. With this lateral closure, the preoral orifice became the only continuity between the complex of airspaces (‘hollows’) inside the head and the air outside the body.

Diagnosis of the identities and evolutionary origins of the lateral head structures involved in this closure has gone through a tortuous history (Spooner, 1938:16; Parsons, 1974), with no resolution, and the most appreciable explanation for this is that they were cell biological events that probably occurred in different