Entomol. exp. appl. 51: 155-162, 1989. �9 1989 Kluwer Academic Publishers. Printed in Belgium. 155

Host plant resistance in Brachiaria grasses to the spittlebug Zulia colombiana

A. Ferrufino* & S.L. Lapointe Centro Internacional de Agricultura Tropical (CIA T), A.A. 6713, Carl, Colombia; * Current address: Instituto Boliviano de Teenologia Agropecuaria (IB TA), Casilla # 2395, Cochabamba, Bolivia

Accepted: February 9, 1989

Key words: Host plant resistance, antibiosis, tolerance, Cercopidae, Zulia colombiana, Brachiaria, Andropogon

Abstract

Twelve forage grass accessions including 11 accessions of Brachiaria Griseb, were evaluated in a glasshouse for host plant resistance to nymphs and tolerance to feeding damage caused by adults of Zurla colombiana (Lallemand) (Homoptera: Cercopidae). Resistance to nymphs was evaluated with a technique that provided uniform environmental conditions and abundant feeding sites. B. brizantha Stapf (cv. Marandfi) was the most resistant of the accessions tested based on nymphal mortality, duration of nymphal stadia, and weight of adult females. Andropogon gayanus Kunth, resistant to spittlebug attack in the field, was susceptible under the conditions of this study. While growth habit and rooting charac- teristics may contribute to field resistance, other resistance factors are present within the genus Brachiaria, particularly in the case of B. brizantha cv. Marandfi. The number of insect-days causing severe damage in the most tolerant species (B. dictyoneura Stapf and B. humidicola Schweick) was approximately six times greater than that necessary to cause the same level of damage to the most susceptible species (B. ruziziensis Germain & Evrard and B. decumbens Stapf). No difference was found in regrowth capacity between infested and noninfested plants within accessions. There was a significant positive correlation between number of insect-days causing severe damage (tolerance) and regrowth of infested plants.

Introduction

Species of the grass genus Brachiaria have been extensively planted as forage in the American tropics due to their excellent productivity and adaptation to soils of low fertility and low pH. It is estimated that more than 15 million hectares are currently planted to Brachiaria spp. (Toledo & Nores, 1986). A limiting factor in productivity and persistance of these grasses is their susceptibility to damage caused by various species of spittle- bugs (Cercopidae). Chemical control methods are

uneconomical due to the low value per unit area of pastures under extensive grazing systems. Host plant resistance, however, has a great potential for controlling pests in tropical forrages in the context of an integrated, low-input, environmentally appropriate program (Lenn6 et al., 1980).

Spittlebug pests of pasture grasses are capable of causing significant damage through removal of sap from xylem elements thereby contributing to water deficit and a general weakening of the plant (Naves, 1980). However, damage caused in forage grasses by adult spittlebugs such as Zurla colom-

156

biana, Aeneolamia reducta (Lallemand), and others is more evident. The initial symptom after adult feeding is a watery and chlorotic spot around the feeding site. Chlorotic lesions expand longitudinally and eventually become necrotic, apparently due to toxic factors contained in the insect saliva and injected into the plant during feeding (Byers & Wells, 1966; Val6rio, 1985). This damage is commonly referred to as froghopper blight. Damage caused by adult spittlebugs has been related to duration of feeding and number of insects feeding per plant (Byers & Wells, 1966). Nymphs feed on exposed roots as early instars and move to the base of stems as they develop. Although nymphs are not known to possess a salivary toxin, they are capable of causing a generalized chlorosis of the plant when present in large numbers, presumable due to removal of water from the xylem.

Among species of tropical pasture grasses, there is considerable variation in amount of visi- ble foliar damage caused by a given insect density, perhaps as a result of tolerance to toxins con- tained in spittlebug saliva. The dramatic failure in Brazil of B. decumbens due to its high suscepti- bility to feeding damage by Deois incompleta (Walker) and Zulia entreriana (Berg) and recent spread of the tolerant B. humidicola serves as an example of the potential value of tolerance to forage grass production in tropical lowlands of Latin America (Toledo & Nores, 1986; Silva, 1982). However, recent experience in Brazil indi- cates that B. humidicola is now showing severe damage from spittlebug attack perhaps as a result of increased insect populations that this tolerant variety is able to support.

Recent acquisition of a large collection from Africa now makes it possible to expand the genetic diversity ofBrachiaria spp. available to growers in the New World tropics. In order to properly select and characterize this new material, an adequate screening methodology for categories of host plant resistance is needed. The purpose of this study was to develop a method for rapid selection under glasshouse conditions of germplasm with antiobiotic resistance to spittlebugs and to char- acterize tolerance in promising tropical forage

species to damage caused by adults of Z. colom- biana in preparation for a large scale screening of new Brachiaria accessions.

Materials and methods

Antibiosis. The study was conducted in a glasshouse at Palmira, Colombia. Eleven plants each of 12 accessions were propagated vegeta- tively from field plots. Andropogon gayanus CIAT 621 (cv. Carimagua 1) is an erect, tufted perennial grass reported to be resistant to spittlebug in the field (Cosenza, 1982). B. decumbens was one of the first species of Brachiaria widely planted in South America and is characterized by a decum- bent, stoloniferous growth habit, rooting from the lower nodes, and susceptibility to damage caused by adult spittlebugs. We selected B. decumbens CIAT 606 (cv. Basilisk) as a susceptible check and B. decumbens CIAT 6132, another high yield- ing accession well adapted to acid soils. B. ruziziensis (CIAT 654 and CIAT 6419) is a prostrate type and highly susceptible to spittlebug damage in the field. B. brizantha (CIAT 665 and CIAT 6294 cv. Marandfi) is also tufted, although not as pronounced as A. gayanus, with suberect stems. B. brizantha and B. decumbens intergrade morphologically although, according to Lock (1977), they differ sufficiently to justify their status as separate species. B. humidicola (CIAT 675, CIAT 6369, CIAT 6705, and CIAT 6707) is char- acterized by narrow leaves, strongly stoloniferous and prostrate growth habit, tolerance to spittlebug damage, and lower nutritional quality than B. decumbens. B. dictyoneura CIAT 6133 is very similar morphologically to B. humidicola but with a slightly tufted growth habit.

Accessions were planted in 7 inch (17.8 cm) plastic pots filled with sterilized soil (Ultisol, pH 4.8) and fertilized at establishment with the equivalent of 50 kgha- 1 each of N, P, and K. One month after planting, nitrogen (urea) was applied at the rate of 50 kg ha- 1.

At planting (approximately eight weeks before infestation), plots were covered with aluminum foil with a central opening for the grass stems in

order to stimulate proliferation of secondary roots at the soil surface. In this way, abundant feeding sites were provided for spittlebug nymphs.

Eggs of Z. colombiana were obtained from adults collected in the field at Santander de Quilichao, Cauca department, Colombia. Adults were placed in an oviposition cage (50 x 50 x 80 cm) provided with a tray of finely sieved, moist soil into which the females ovi- posited. Eggs were separated from the soil by passing through a series of sieves (42, 60 and 120 mesh). Eggs were disinfected in 0.5~o sodium hypochlorite for 5 minutes, washed in distilled water, and incubated in petri dishes on moist filter paper until fully developed (approximately 15 days). Thirty fully developed eggs were placed at the base of each plant on moist filter paper and allowed to eclose. Three days after infestation, the filter paper was removed along with those eggs that did not eclose. The initial mean number of nymphs per accession varied from 9.5 to 13.2 nymphs per plant.

Nymphs were examined daily until the final moult to determine length of stadia and survival. When nymphs reached fifth instar, the aluminum foil was removed and an organdy mesh cage was placed over each plant to collect emerging adults. Adults were collected daily, sexed, dried in an oven, and weighed.

Infested plants were arranged in a glasshouse in a randomized block design with 11 replicates. During the experimental period temperature in the glasshouse fluctuated in synchrony with photo- period between 19 and 27 ~ with a mean of 23.2 + 1.2 o C. Relative humidity ranged between 60 ~o and 90 ~o with a mean of 82 ~o. Temperature under the aluminum foil inside three pots chosen at random was measured thrice daily with a fan operated psychrometer over the experimental period at 7 : 30 AM, 1 PM, and 4 PM with three repetitions. Mean temperature was 23.9 + 2.5 ~ C and mean relative humidity was 85.8 _+ 6.1~o.

Data were analyzed by ANOVA and Duncan's Multiple Range test. Percent survival was ana- lyzed using the natural log transformation sug- gested for percentages by Steel & Torrie (1985). Tolerance. Upon completion of the antibiosis trial,

157

plants were cut to a height of 5 cm above the root crown, and fertilized with the equivalent of 50 kg ha 1 each of N, P, and K. Two weeks later, a second application of nitrogen (urea) was applied at the rate of 50 kg h a - i. Plants were allowed to recover for 5 weeks, covered with individual organdy mesh cages and infested with adults. The study was conducted in a glasshouse, as above. During the experimental period, temperature in the glasshouse fluctuated in synchrony with photoperiod between 19 and 27 ~ with a mean of 24.3 + 0.9 ~ Relative humidity ranged between 60~o and 90~o with a mean of 82~/o. Adults were obtained from a laboratory colony of Z. colombiana maintained in a glasshouse at CIAT, Palmira. Due to the large number of insects required and limited availability of insects from the colony, initial infestation was 4 adults per plant and was increased by 2 adults per plant every 5 days until severe feeding damage was observed. In the case of the most tolerant acces- sion, B. humidicola CIAT 675, a maximum infes- tation of 22 insects per plant was obtained. All adults were a maximum of one day old when placed on caged plants and dead insects were replaced daily with one-day-old adults. Plants were arranged on tables in a completely ran- domized design with six replications and two uninfested pots per accession as controls.

Damage was scored daily on a visual scale of percent foliar area affected as follows: 1. No damage 0 ~o foliar area affected. 2. Slight danaage 20-40 ~o foliar area affected. 3. Moderate damage 40-60 ~o foliar area affected. 4. Severe damage 60-80 ~ foliar area affected.

When severe feeding damage was observed, adults were removed and foliage was cut, dried in an oven at 65 ~ and weighed. Plants were allowed to regrow for 40 days and production of dry matter was again measured and compared with uninfested controls.

Tolerance was measured in units of insect-days (product of the number of adult spittlebugs and number of days at that level of infestation, summed over the period of infestation) required to achieve severe feeding damage. Mean daily adult mortality was compared by ANOVA when the

158

most susceptible accession showed severe dam- age.

Results

Antibiosis. Egg eclosion and early instar survival were high on all accessions. Figure 1 shows survivorship by instar for 5 accessions that repre- sent the range of response obtained. In general, first and second instar survival was high and greatest mortality occurred during third and fourth instars.

With the exception of B. decumbens, survival was lowest on B. brizantha CIAT 6294 (cv. Marandfl) (Table 1). In general, nymphs reared on Marandti produced small spittle masses and migrated frequently within the pots. In contrast, nymphs reared on B. dictyoneura, B. humidicola, and A. gayanus produced abundant spittle and did not migrate. Nymphs reared on Marandfl developed more slowly than those on other acces- sions and emerged as adults on the average 14 days later than those on B. dictyoneura CIAT 6133.

Adult females reared on Marandfi weighed on the average 24~o less than those emerging from B. ruziziensis CIAT 6419 (Table 1). Correlations

i

.m

1oo

90

80

7 0

5 0 " --0- B decurnbens CIAT606 B bMzan'(ha CIAT 6294

40

Instar

Fig. 1. Stage-specific survivorship of Z. colombiana nymphs reared on five grass species in a glasshouse.

were highly significant between nymphal survival to adult and duration of nymphal stadia (r 2 = 0.713, P = 0.0003) (Fig. 2) and between adult female weight at emergence and duration of nymphal stadia (r 2 = 0.677, P = 0.0006) (Fig. 2). The correlation between nymphal survival to adult and adult female weight at emergence was not significant (r 2 = 0.272, P -- 0.068).

Tolerance. There was a positive correlation (r 2 = 0.77, P < 0.01)between biomass (grams dry weight/plant) and days of infestation required to

Table 1. Vital statistics of Z. colombiana reared on various tropical forage grasses

Species Accession Survival N Duration of to adult nymphal stadia (%)* (days)*

N Weight of female adults (mg)*

N

B. humidicola 6707 95.9 a 145 46.3 ab 140 B. dictyoneura 6133 95.7 a 141 44.2 a 135 B. humidicola 6705 94.0 a 138 48.6 ab 133 B. brizantha 665 93.6 a 118 48.2 ab 103 B. humidicola 675 92.9 a 130 45.9 ab 125 B. humidieola 6369 88.3 a 122 48.7 ab 111 B. ruziziensis 654 87.1 a 125 45.9 ab 114 A. gayanus 621 86.6 a 107 46.1 ab 93 B. decumbens 6132 83.0 ab 140 46.6 ab 122 B. ruziziensis 6419 74.5 ab 118 46.7 ab 89 B. decumbens 606 63.7 bc 106 51.6 b 67 B. brizantha 6294 47.3 c 105 58.3 c 51

14.5 abc 14.9 ab 12.8 cd 12.6 cd 14.2 abc 13.8 abc 14.0 abc 13.9 abc 13.4 abc 15.2 a 12.9 cd 11.5 d

68 73 65 46 69 59 45 48 61 42 25 24

* Means followed by the same letter do not differ (c~ = 0.05, Duncan's Multiple Range Test).

I I

I I L I

=~

I 0 0 A

90

80

70

60

50

40

' I I I I ' ~ 1 I ' L '

B

] ~ . y= 24 84- 0 23X 14 J ~11 R = -0 82**

,-~ ~ ~

_~ 13- ~ m m @~E ~ . II

I 40 50 60

D u r a t i o n o f n y m p h a l s t a d i a ( d a y s )

Fig. 2. Correlation between duration of nymphal instars and A) survival of nymphs to adult and B) adult female weight at emergence for Z. colombiana reared on 12 grass accessions.

40 ' l , I , I , I , I

y=-030 +275X / *

30 R2 = 0.775 * ~ / /

2O

10 . . . . . 4 6 8 I0 12 14 6

Dry w e i g h t (g /p lant )

Fig. 3. Correlation between plant biomass (dry weight) and tolerance to adult spittlebug damage as measured by days of infestation necessary to cause severe foliar damage to plants grown in a glasshouse.

159

B ru~izler~Is 654

B IV2.1zmI~Is 6419 B decumben~ 6132

B decumbens 606

B blJzantha 655

B hu~Ico]a 6369

B hu~icola 6707

A ~aya~11~ 621

B brlzalxtAa 6294

B dlctyone'd~ 6133

B humJdlcola 6705

B hwnldlCOla 675

I

I I

100

I

I

200 300 Insect-days

400 I

500

Fig. 4. Meannumber of insect-days resulting in severe follar damage by nymphs of Z. colombiana to 12 grass accessions in the greenhouse. Bars are standard deviation.

cause severe feeding damage (Fig. 3). Therefore, biomass was used as a covariable in the analysis of means by ANOVA and Duncan's Multiple Range Test. The number of insect-days causing severe damage in the most tolerant accessions (B. humidicola CIAT 675, 6705, and B. dictyo- neura CIAT 6133) was between five and six times greater than that necessary to cause the same damage to the most susceptible accessions (B. ruziziensis CIAT 654 and 6419, B. decumbens CIAT 6132) (Fig. 4).

No difference was found (c~ = 0.05, ANOVA) in regrowth capacity between infested and non- infested plants. However, there was a significant positive correlation between number of insect- days causing severe damage and regrowth (r 2 = 0.69, P = 0.0009). No significant difference was found in daily adult mortality (c~ = 0.05, ANOVA).

Discussion

Survival of first and second instar nymphs in this study was high for all accessions studied. Other authors have reported high early instar mortality (Magalhaes, 1982; Barrientos, 1984; Nilakhe etal . , 1985). It is possible that low relative humidity and poor availability of feeding sites had a large effect on survival of early instar nymphs. In this study, we assume that such mortality fac- tors were largely excluded due to near optimal

160

conditions of relative humidity and abundance of superficial rootlets afforded by the aluminum foil cover.

It is interesting to note the high survival of nymphs reared on A. gayanus. Under field condi- tions, this grass is seldom infested with spittlebugs and was therefore selected as a resistant check. Various authors have reported antibiosis as the category of resistance in A. gayanus (Cosenza, 1982; CIAT, 1982; Lenn6 & Calderdn, in press). Lenn6 & Calder6n (in press) suggest that cate- chins and cyanogenic glycosides in leaves, stems, and roots ofA. gayanus are related to an antibiosis type of resistance to spittlebug. No study to date, however, has demonstrated a relationship between secondary compounds in A. gayanus and insect response nor that such chemicals are present in the xylem, principal feeding site of spittlebug nymphs and adults. The results of this study suggest that resistance exhibited by A. gayanus in the field may not have a chemical basis. A more likely explanation is that under field conditions, the growth habit ofA. gayanus (deep- rooting, tufted type) does not provide feeding sites and microclimatic conditions adequate for devel- opment of spittlebug nymphs. It is important to note that the aluminum foil cover used in this study stimulated proliferation of superficial rootlets and provided uniform conditions of tem- perature, light, and relative humidity. Suscepti- bility to Z. colombiana under these conditions suggests that secondary chemistry is not operating as a resistance mechanism in A. gayanus.

Growth habit and its influence on micro- climatic factors near the soil surface may be an important character for selection of resistant germplasm and may be a significant source of mortality for early instar nymphs when they are most susceptible to dessication. However, it appears that additional resistance factors are present within the genus Brachiaria. Marandfi is also resistant to spittlebugs in the filed (Cosenza, 1982). Unlike A. gayanus, however, Marandfi retained its relatively high level of antibiosis under the conditions of this study. High nymphal mortality has also been observed for Z. entreriana (Berg) in Brazil on Marandfi by Nilakhe et al.

(1985). In this study, microclimatic differences that certainly exist in the field between grasses with widely different growth habits were mini- mized. Increased nymphal mortality, prolonged development time, and reduced adult female weight of Z. colombiana reared on Marandfi may be attributed to plant defenses other than growth habit, including physical defenses, allelochemi- cals, or nutritional factors. The mechanism of resistance will be of particular interest if a breeding program for Brachiaria is initiated.

The high mortality of nymphs on B. decumbens CIAT 606 may not be related to an antibiotic plant defense mechanism. Plants of this accession suffered an accentuated chlorosis during the period of infestation. Deterioration of B. decum- bens probably contributed to nymphal mortality and is an indication of the high susceptibility of this species to damage caused by spittlebug nymphs. Future studies of antibiosis should employ lower levels of infestation on such highly susceptible accessions to eliminate mortality due to competition.

Taliaferro et al. (1969) suggested that tolerance to spittlebug in Cynodon dactylon (Coastal Bermudagrass) was due to either preferential feeding or tolerance to salivary toxin. Although adult feeding was not directly measured in this study, daily adult mortality on caged plants did not differ by accession. Regrowth capacity of individual accessions was unaffected by the ex- perimental infestations but tolerance was corre- lated with plant regrowth capacity.

A wide range of tolerance exists within acces- sions of Brachiaria. The danger inherent in select- ing germplasm solely on the basis of tolerance has been demonstrated by the case ofB. humidicola in the humid tropics of Brazil. However, tolerant materials such as B. humidicola and B. dictyoneura will be useful where insect populations are maintained below an economic threshold by biotic or abiotic factors. In areas where potential for spittlebug population growth is high, an ideal cultivar would be one that possesses antibiotic properties acting to suppress insect populations and tolerance to insect feeding should insect populations become significant. Results pres-

ented here suggest that Marandfi is such a grass. It is the most resistant accession of Brachiaria tested to date and possesses a moderate degree of tolerance to adult feeding damage. Using evalua- tion techniques developed here together with field evaluations of edaphic adaptation and response to natural spittlebug infestations, we hope to select accessions adapted to acid, infertile soils of the lowland American tropics and resistant to spittlebug. The limited amount of germplasm that has been introduced to date suggests that there is as yet a large potential for selecting valuable mate- rials from naturally occurring diversity with eda- phic adaptation as well as pest and disease resistance.

The methodology described for rearing nymphs provides a practical system for evaluation of host plant resistance in Brachiaria to spittlebugs. At CIAT, we are now rearing Z. colombiana and A. reducta using this technique. Current efforts include a comparison of these and other spittle- bugs to determine variability of response to resistant materials within the complex of Latin America species. The high survival and rapid development of Z. colombiana nymphs on B. dictyoneura and B. humidicola recommend their use as susceptible checks in future screening and for spittlebug mass-rearing.

R6sum6

ROsistance de Brachiaria aux dOgdts des adultes du Cercopidae, Zulia colombiana

Brachiaria est une gramin6e fourrag6re promet- teuse pour les sols tropicaux acides, satur6s d'aluminium. Z. coIombiana est un Cercopidae tr~s r6pandu, limitant l'utilisation de Brachiaria en Am6rique Latine. La r6sistance (antibiose et tol6rance) i~ Z. colombiana, de Brachiaria d'origi- nes diverses a 6t6 examin6e. B. brizantha cv. Marandfl s'est r6v616 le plus r6sistant d'apr~s la forte mortalit6 larvaire, la prolongation du d6vel- oppement larvaire, et le poids r~duit des femelles adultes de Z. colombiana. Andropogon gayanus, r6sistant dans la nature, s'est r6v616 sensible. Ces

161

r6sultats sugg6rent que cette r6sistance de A. gayanus dans la nature pourrait atre due fi la structure du v6g6tal et/l son mode de croissance. Dans le cas de B. brizantha cv. Marandfl, des facteurs suppl6mentaires de r6sistance, mis en 6vidence ~i partir de diff6rents modes de crois- sance, ont 6t6 61imin6s, de fagon/t identifier les mdcanismes de l'antibiose pr6sents chez Brachiaria. Une grande gamme de r6sistance aux attaques alimentaires a 6t6 observ6e chez Brachiaria. Les plus r6sistants ont besoin de 6 fois plus de jours d'attaque par Z. colombiana pour provoquer les d6ggtts observ6s sur individus sensi- bles.

Acknowledgement

We thank Dr. Elkin Bustamante (CATIE, Costa Rica) for suggestions and advice, Myriam Cristina Duque for statistical consultation, and the Ento- mology section of the Tropical Pastures Program of CIAT for technical assistance.

References

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Byers, R.A. & H.D. Wells, 1966. Phytotoxemia of coastal bermudagrass caused by two-lined splttlebug Prosapia bieincta (Homoptera: Cereopidae). Ann. Entomol. Soc. Amer. 59(6): 1067-1071.

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Loch, D.S., 1977. Brachiaria decumbens (signal grass) - a review with particular reference to Australia. Trop. Grasslands 11 : 141-157.

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Val6rio, J.R., 1985. Caraterizaggo e avaliaggo do dano causado pelo adulto da cigarrinhadas-pastagens Zulia entreriana (Berg, 1879)em Brachiaria decumbens Stapf. cv. Basilisk. Tesis Doutor em Ciencias, Piracicaba, Brasil, Universidade de $5o Paulo. 152 p.