The use of entomopathogenic nematodes to control citus mealybug, Planococcus citri (Hemiptera: Pseudococcidae) on citrus in South Africa

Van Niekerk, Sonnica (2012-03)

Thesis (MScAgric)--Stellenbosch University, 2012.


ENGLISH ABSTRACT: Planococcus citri (Risso) (Hemiptera: Pseudococcidae), the citrus mealybug, is a highly destructive pest of citrus in South Africa. The ability of mealybugs to develop resistance to chemicals, as well as their cryptic nature and protective wax coverings, all individually and combined, impair the ability of insecticides to control them. Furthermore, chemicals deplete natural enemy populations that usually establish control over mealybug populations well before harvest in March to May. The inability of chemicals to control P. citri successfully and the growing public awareness of the detrimental environmental effects and health risks that are associated with pesticides are pressuring citrus growers to find alternative methods to chemical control. Entomopathogenic nematodes of the Rhabditida order, belonging either to the families Heterorhabditidae or Steinernematidae, have proven to be valuable biocontrol agents of a variety of insect pest species. These nematodes are, however, not yet commercially available in South Africa. Various laboratory bioassays were conducted to determine the potential of entomopathogenic nematodes to control P. citri. Adult female P. citri were screened for susceptibility to six indigenous nematode species. Planococcus citri was found to be most susceptible to Steinernema yirgalemense and Heterorhabditis zealandica, causing 97% and 91% mortality, respectively. Both H. zealandica and S. yirgalemense completed their life cycles inside adult female P. citri. Under optimal conditions, H. zealandica and S. yirgalemense, at an application rate of 170 infective juveniles (IJs) / insect (LD90), were able to control P. citri efficiently with a discriminating dosage of 11(LD50). The water activity (aw) bioassay indicated that S. yirgalemense was two times more tolerant to lower levels of free water than H. zealandica, with aw50 = 0.96 and aw90 = 0.99, compared to H. zealandica with aw50 = 0.98 and aw90 = 1.01. Furthermore, S. yirgalemense proved able to locate and infect P. citri at a faster rate than H. zealandica. Nematode activity was not significantly affected when exposed to 15°C, 20°C and 25°C. Results of the exposure trial also showed that the first 2−4 h post-application was the most decisive time for establishing successful infection of mealybugs. Further experiments were conducted to determine the ability of two polymer products, Zeba® and Xanthan gum, and a surfactant, Nu-Film-P®, to improve nematode application suspensions. Despite not being as effective as Xanthan gum, Zeba® still retarded sedimentation significantly. The addition of 0.02% Xanthan gum did not improve the ability of nematodes to control P. citri at 60% and 80% relative humidity containing H. zealandica and 0.03% Zeba® increased mortality by 14% at 60% RH and by 22% at 80% RH. The same polymer formulation was tested with S. yirgalemense, with mortality being found to have increased by 21% at 60% RH and by 27% at 80% RH. The combined addition of Nu-Film-P® and Zeba® was able to retard sedimentation significantly, increasing the average number of nematodes deposited on 2-cm2 leaf discs. To illustrate those factors that should be taken into consideration before applying nematodes in an integrated pest management (IPM) programme for citrus, the compatibility of H. zealandica and S. yirgalemense with biological control agents and agrochemicals to which they are most likely to be exposed was determined. Bioassays showed the coccinellid beetle, Cryptolaemus montrouzieri, to be susceptible to both nematode species. Beetle larvae proved to be highly susceptible, with H. zealandica obtaining 80%, and S. yirgalemense obtaining 92% control. Adult beetles were found to be twice as susceptible to S. yirgalemense, with 64% mortality recorded, as to H. zealandica, with 30% mortality recorded. Tolerance of H. zealandica and S. yirgalemense infective juveniles to the affect of aqueous solutions of an insecticide (Cyperphos 500 E.C.®), two biopesticides (CryptogranTM and HelicovirTM), and two adjuvants (Nu-Film-P® and Zeba®) on the infectivity and survival of H. zealandica and S. yirgalemense was evaluated. Heterorhabditis zealandica proved to be highly compatible with all products tested. Significant increase in mortality of S. yirgalemense was recorded after 12 h exposure to CryptogranTM, HelicovirTM and Cyperphos 500 E.C.®, and after 6 h exposure to Nu-Film-P®. However, there was no decrease in the ability of S. yirgalemense to infect the insect hosts. Towards selecting the appropriate nematode species for field studies, the ability of S. yirgalemense and H. zealandica to control P. citri was first evaluated under less harsh simulated glasshouse conditions. The ability of both the above-mentioned nematode species to control P. citri, with and without the addition of 0.03% Zeba® and 0.06% Nu-Film-P®, was evaluated in a growth chamber at 75 ± 8% relative humidity and 22°C. All treatments resulted in significantly higher mortality of adult female P. citri than did the control. The addition of 0.03% Zeba® and 0.06% Nu-Film-P® to an H. zealandica and an S. yirgalemense suspension increased resultant mortality from 26% to 30%, and from 34% to 45%, respectively. This increase in mortality was, however, not significant. The ability of this formulation to prolong the ability of S. yirgalemense to infect P. citri and to prolong nematode survival was also evaluated under the same conditions and showed the formulation to improve both infectivity and survival for up to 2–3 h post-application. In the semi-field trial, S. yirgalemense was unable to obtain significant control of P. citri without the addition of 0.03% Zeba® to nematode application suspensions. However, with the addition of 0.03% Zeba® S. yirgalemnse was able to obtain up to 53% control. The study established that the polymer product Zeba® improves the ability of S. yirgalemense to infect P. citri by retarding desiccation and by buffering nematodes from suboptimal environmental conditions.

AFRIKAANSE OPSOMMING: Planococcus citri (Risso) (Hemiptera: Pseudococcidae), die sitrus witluis, is ʼn baie skadelike pes van sitrus in Suid Afrika. Die vermoë van witluise om weerstand teen chemiese middels te ontwikkel, hul kriptiese lewenswyse en die beskermende waslaag wat hul liggame omhul inhibeer gesamentlik en individueel die vermoë van insektisiede om witluis bevolkings te beheer. Chemiese middels verminder ook die natuurlike vyande wat gewoonlik witluis bevolkings beheer voor die sitrus oestydperk in Maart tot Mei. Die onvermoë van chemiese middels om P. citri suksesvol te beheer en verhoogde bewustheid van die publiek rond om die vernietigende omgewings impak en gesondheidsrisiko’s verbonde aan chemiese insek beheer, noodsaak sitrus produsente om alternatiewe beheermetodes te ontwikkel. Entomopatogeniese nematodes, van die orde Rhabditida wat aan die families Heterorhabditidae of Steinernematidae behoort, is bekend as effektiewe biologiese beheeragente van ʼn verskeidenheid insek pes spesies. Hierdie nematodes is egter tans nie kommersieel beskikbaar in Suid- Afrika nie. Om ten einde die vermoë van plaaslike nematode spesies te bepaal om P. citri te beheer, is verskeie biotoetse in die laboratorium uitgevoer. ʼn Vinnige siftings proses is uitgevoer om vas te stel watter plaaslike nematode spesies die hoogste persentasie mortaliteit van P. citri wyfies veroorsaak. Daar is bevind dat P. citri die mees vatbaarste is vir Steinernema yirgalemense en Heterorhabditis zealandica wat 97% en 91% mortaliteit respektiewelik veroorsaak het. Die ontwikkeling van beide H. zealandica en S. yirgalemense na die infektering van volwasse P. citri wyfies is gevolg en daar is bevind dat beide nematode spesies hul lewensiklusse kon voltooi in insek kadawers. Onder optimale toestande was H. zealandica en S. yirgalemense in staat om P. citri effektief te beheer, as hul teen ʼn konsentrasie van 170 infektiewe larwes (JIs) per insek (LD90) toegedien word, met ʼn diskriminerende toedienings dosis van 11 (LD50). ʼn Water aktiwiteit biotoets het gewys dat S. yirgalemense twee keer so verdraagsaam is teenoor laer vlakke van vrye water as wat H. zealandica is met aw50 = 0.96 en aw90 = 0.99, in vergelyking met H. zealandica met aw50 = 0.98 en aw90 = 1.01. Resultate van ʼn blootstellings toets het ook gewys dat S. yirgalemense die vermoë het om P. citri vinniger op te spoor en te infekteer as H. zealandica. Die blootstellings toets was uitgevoer teen 15°C, 20°C en 25°C en daar is bevind dat die aktiwiteit van H. zealandica nie beduidend verhoog het met ʼn verhoging in temperatuur nie. Resultate vir die blootstellings toets het ook gewys dat die eerste 2 tot 4 uur na toediening van nematodes die mees kritieke tyd is om suksesvolle infektering van witluise te bevestig. Verdere eksperimente is uitgevoer om te bepaal of die byvoeging van twee polimeer produkte nl. Zeba® en Xanthan gum en ʼn benatter Nu-Film-P®, nematode toedienings suspensies kan verbeter. Deur 0.02% Xanthan gum by nematode toedienings suspensies te voeg is die vermoë van nematodes om P. citri by 60% en 80% relatiewe humiditeit (RH) te beheer nie beduidend verhoog nie, terwyl die byvoeging van 0.03% Zeba® wel ʼn beduidende verhoging in mortaliteit veroorsaak het. ʼn Suspensie van H. zealandica en 0.03% Zeba® het beheer met 14% by 60% RH en met 22% by 80% RH verhoog. Die invloed van dieselfde polimeer formulasie was ook getoets op S. yirgalemense en mortaliteit het verhoog met 21% by 60% RH en met 27% by 80% RH. Ten spyte daarvan dat “Xanthan gum” nie so effektief was om afsakking van nematodes te verhoed nie, het die byvoeginging van 0.03% Zeba® by toedienings suspensies steeds ʼn beduidende invloed gehad. Die gekombineerde byvoeging Nu-Film-P® en Zeba® was in staat om die gemiddelde aantal nematodes gedeponeer op 2-cm2 blaar skyfies te verhoog. Om te wys watter faktore in ag geneem moet word voor EPNs as deel van ʼn geïntegreerde pes beheer program toegedien word, is die verenigbaarheid van H. zealandica en S. yirgalemense vir biologiese beheer agente en landbouchemikalieë, waaraan hulle heel waarskynlik blootgestel gaan word, bepaal. Biotoetse het gewys dat die liewenheersbesie, Cryptolaemus montrouzieri, vatbaar is vir beide nematode spesies. Die larwale fase was hoogs vatbaar met mortaliteit van 80% en 92% verkry deur die toedeining van H. zealandica en S. yirgalemense respektiewelik. Daar is ook bepaal dat volwasse besies twee maal so vatbaar is vir S. yirgalemense wat 64% mortaliteit veroorsaak het, as vir H. zealandica met 30% mortaliteit aangeteken. Verdraagsaamheid van IJs van H. zealandica en S. yirgalemense met oplossings van ʼn insektisied (Cyperphos 500 E.C.®), twee bio-plaagdoders (CryptogranTM en HelicovirTM), en twee byvoegmiddels (Nu-Film-P® en Zeba®), vir infektiwiteit en oorlewing is bepaal. Resultate het gewys dat H. zealandica hoogs verenigbaar is met alle produkte wat getoets is. ʼn Beduidende verhoging in mortaliteit van S. yirgalemense is aangeteken na 12 ure se blootstelling aan CryptogranTM, HelicovirTM en Cyperphos 500 E.C.®, en na 6 ure se blootstelling aan Nu-Film-P®. Resultate het egter ook gewys dat die vermoë van S. yirgalemense om om gashere te infekteer nie deur beinvloed word nie. Om ten einde te bepaal watter een van H. zealandica of S. yirgalemense die beter isolaat sou wees om te gebruik in veldproewe, is hul vermoë om P. citri te beheer eers evalueer onder minder ongunstige gesimuleerde glashuis toestande. Die vermoë van beide nematode spesies om P. citri te beheer is bepaal in ʼn groeikamer by 75 ± 8% (RH) met en sonder die byvoeging van 0.03% Zeba® en 0.06% Nu-Film-P®. Mortaliteit verkry deur alle behandelings was beduidend hoër as die kontrole. Deur 0.03% Zeba® en 0.06% Nu-Film-P® by toedienings suspensies van H. zealandica en S. yirgalemense te voeg is die gevolglike mortaliteit van P. citri verhoog van 26% na 30% en van 34% na 45%, respektiewelik. Hierdie verhoging in mortaliteit was egter nie beduidend nie. Die vermoë van die bo-genoemde formulasie om oorlewing van S. yirgalemense en infeksie tydperk van P. citri te verleng was bepaal onder dieselfde toestande en daar is gevind dat die formulasie beide die infektiwiteit en oorlewing van nematodes met 2 tot 3 uur na toediening verleng het. Tydens die semi-veldproef was S. yirgalemense nie in staat om beduidend hoër mortaliteit van P. citri te verkry, in vergelyking met die kontrole, sonder die byvoeging van 0.03% Zeba® by toedienings suspensies nie. Met die byvoeging van Zeba® is daar tot 53% beheer verkry. Die studie het onweerlegbaar gewys dat die polimeer produk Zeba® die vermoë van S. yirgalemense om P. citri te infekteer verbeter deur die uitdroging van nematodes te vertraag en deur hul te beskerm teen ongunstige omgewings toestande.

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