TABLE CONTENTS

Two ~ 400 bp fragments of rRNA loci flanked by “universal” primer pairs (NF1 - 1573R 18S
and D3A - D3B 28S) were used to identify nematode taxa. We hand-picked 44 known
nematode species in known frequencies to set up 4 artificial metagenomic samples. Two
metagenomic samples consisted of DNA of all nematode species extracted and amplified
together (18S mPCR and 28S mPCR). Two additional samples came from pooling PCR
products from single nematode species PCR reactions (18S sPCR and 28S sPCR). All
metagenomic samples were A-Amplicon sequenced on GS FLX and run separately on a 16-
chambered plate. The total number of reads ranged from 4159 to 14771 per sample. Out of all
reads, ~82% comprised reads of at least 200 bp and within those reads ~91% were identified
as matching our referenced 44 species. While ~52% of reads gave 100% identity match,
about 30% of reads varied by 1-2 bp, and the rest of the reads varied by >3 bp. Although
neither diagnostic region recovered all nematode species, 18S resulted in higher species
recovery (~90%) than 28S barcode (~80%) and the use of both barcodes improved the
detection level of nematode species. Of the >199 bp reads that were unmatched to our
nematode database (~8%), the majority were identified as fungal and ~0.4% were identified
as chimeras. The frequency distribution of reads did not mirror the frequency distribution of
nematode species.
Overall, results strongly support the suitability of 454 technology for identification of all
nematode individuals from environmental samples. At this point, however, the use of the
distribution reads for inferring the relative abundances of species within a nematode
community is premature.
Effects of Cadmium, Lead and Zinc on the Entomopathogenic Nematode
Steinernema feltiae
Kelly, E. & T. Kakouli-Duarte
Molecular Ecology and Nematode Research Group, Department of Science and Health, Institute of Technology
Carlow, Kilkenny Road, Carlow, Ireland
Nematodes have been well documented as useful bioindicator organisms for environmental
monitoring. This project investigates the suitability of the entomopathogenic nematode
Steinernema feltiae as a sentinel species of heavy metal pollution in areas of mining activity
in Ireland, where in recent years, public health concerns were raised. The nematode is widely
occurring in Ireland and can be easily extracted from soil and cultured in the laboratory. In
addition, the third stage infective juveniles provide suitably age synchronised individuals for
toxicity testing. Experiments were carried out in which the effects of cadmium, lead and zinc
were investigated on several traits of the nematode. Preliminary results revealed that short
term heavy metal exposures of the nematodes did not affect their insecticidal ability against
Galleria mellonella larvae, with the exception the case of lead at the higher dose tested. On
the other hand there were clear negative effects on nematode progeny production, indicating
that fecundity in S. feltiae could be developed as suitable end point for heavy metal toxicity.
The nematodes used in this study belong to a native Irish strain isolated in Bull Island Co.
Dublin. Efforts are on going to isolate S. feltiae from the mining areas of interest so that long
term effects could also be determined. This project endeavours to combine physiological and
behavioural nematode responses with data on genotoxic effects to be used as environmental
risk assessment tools for cadmium, lead and zinc pollution.
5 th International Congress of Nematology, 2008 267