New Zealand is currently the only major fruit producing country in the world that is free of economically important fruit flies. As part of the effort to maintain this status, there is a need to supplement quarantine decision-making procedures with a means of rapidly identifying immature life stage infestations to the species level. Here we describe a molecular method that achieves this, using simple restriction patterns of ribosomal DNA (rDNA) as diagnostic markers. The 18S and 18S plus internal transcribed spacer (ITS) regions were amplified from larval DNA by the polymerase chain reaction (PCR). Nineteen species, spanning four genera (including five subgenera of Bactrocera) were analysed. Restriction analysis of the 18S PCR product provided poor resolution, even at the generic level. Digestion of the 18S + ITS PCR product, however, generated thirteen diagnostic haplotypes as defined by the composite restriction patterns from Rsal, Sau3a Haelll and Alul. No variation was detected at these restriction sites within or between populations. Twenty two restriction enzymes have been screened, but diagnostic RFLPs have yet to been found for six out of the ten Bactrocera (Bactrocera) species; B. passiflorae (Froggatt) cannot be distinguished from B. facialis (Coquillet), nor B. kirki (Froggatt) from B. trilineola (Froggatt) or B. neohumeralis (Hardy) from B. tnjoni (Froggatt). Geographic origin could assist in distinguishing the first four species, but the latter pair are very closely related with overlapping origins, hosts and adult morphology. All six species, however, are considered high risk with respect to their likely establishment in New Zealand. Therefore diagnosis based on this molecular technique would support the same quarantine decision. We consider this method could be useful as a diagnostic technique and discuss directions for further development.

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Sitona discoideus Gylh., an introduced pest of lucerne in New Zealand, exhibits univoltine, aestivatory seasonality in Canterbury and Otago, with each generation appearing in late December. Some two weeks after the December emergence of adults, flights to aestivation sites commenced. These sites were commonly remote from the lucerne stands and included such places as under stones or at the base of trees, fence posts, etc. At the commencement of aestivation, the indirect flight muscles atrophied rapidly, only to redevelop some 6-8 weeks later when autumnal post-aestivatory return flights to lucerne began. During the winter, S. discoideus fed on lucerne foliage and became reproductively mature. By October, age-related adult mortality had begun, and for about two weeks in December no adult weevils were present at all in one Canterbury site. In Otago, there was a distinct generation overlap of 2-3 weeks. The eclosion of new generation weevils was closely synchronised; it is suggested that this may have been the product of differential egg development and egg-laying temperature thresholds and a high fourth instar and pupal develoment threshold. Synchronised emergence of adults tended to accentuate the problem of defoliation caused by their feeding.

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