BACKGROUND:The nematode phylum includes many species key to soil food webs with trophic behaviours extending from feeding on microbes to macrofauna and plant roots. Among these, the plant parasitic cyst nematodes retain their eggs in protective cysts prolonging their survival under harsh conditions. These nematodes, including those from the genus Heterodera, cause significant economic losses in agricultural systems. Understanding of nematode diversity and ecology has expanded through application of genomic research, however, for Heterodera species there are very few available whole genome sequences. Sequencing and assembling Heterodera genomes is challenging due to various technical limitations imposed by the biology of Heterodera. Overcoming these limitations is essential for comprehensive insights into Heterodera parasitic interactions with plants, population studies, and for Australian biosecurity implications. RESULTS:We hereby present draft genomes of six species of which Heterodera australis, H. humuli, H. mani and H. trifolii are presently recorded in Australia and two species, H. avenae and H. filipjevi, currently absent from Australia. The draft genomes were sequenced from genomic DNA isolated from 50 cysts each using an Illumina NovaSeq short read sequencing platform. The data revealed disparity in sequencing yield between species. What was previously identified as H. avenae in Australia using morphological traits is now confirmed as H. australis and may have consequences for wheat breeding programs in Australia that are breeding for resistance to H. avenae. A multigene phylogeny placed the sequenced species into taxonomic phylogenetic perspective. Genomic comparisons within the Avenae species group revealed orthologous gene clusters within the species, emphasising the shared and unique features of the group. The data also revealed the presence of a Wolbachia species, a putative bacterial endosymbiont from Heterodera humuli short read sequencing data. CONCLUSION:Genomic research holds immense significance for agriculture, for understanding pest species diversity and the development of effective management strategies. This study provides insight into Heterodera, cyst nematode genomics and the associated symbionts and this work will serve as a baseline for further genomic analyses in this economically important nematode group.

The potato cyst nematode Globodera rostochiensis originates from the Andean Mountain region in South America and has unintentionally been introduced to all inhabited continents. Several studies have examined the population genetic structure of this pest in various countries by using microsatellite markers. However, merging microsatellite data produced from different laboratories is challenging and can introduce uncertainty when interpreting the results. To overcome this challenge and to explore invasion routes of this pest, we have genotyped 22 G. rostochiensis populations from all continents. Within populations, the highest genetic diversity was observed in the South American populations, the European populations showed an intermediate level of genetic diversity and the remaining populations were the less diverse. This confirmed pre-existing knowledge such as a first introduction event from South America to Europe, but the less diverse populations could originate either from South America or from Europe. At the continental scale, STRUCTURE genetic clustering output indicated that North America and Asia have experienced at least two introduction events. Comparing different evolutionary scenarios, the Approximate Bayesian Computation analysis showed that Europe served as a secondary distribution centre for the invasion of G. rostochiensis into all other continents (North America, Africa, Asia and Oceania).

Hop cyst nematode, Heterodera humuli Filipjev, 1934 was found in soil samples collected from hop farms in Bushy Park, Tasmania and in Merriang, Victoria, Australia. Morphological and molecular characteristics were consistent with those described for this species in other parts of the world. Novel gene sequences of ITS rRNA, 28S rRNA and CO1 mtDNA generated from cysts were compared against sequences of H. humuli available on the NCBI GenBank database, demonstrating little to no genetic variation between Australian hop cyst nematode populations and those from other regions. This report provides the first molecular sequence data for H. humuli from Australia and reports a significant range extension of this cyst nematode from Tasmania to the Australian mainland.

Cyst nematodes of the genus Heterodera are a major group of sedentary plant parasites causing a significant economic impact, restricting production and market access globally (Moens et al. 2018). The ryegrass cyst nematode Heterodera mani is in the Avenae group and is found predominantly in pastures and grasslands in Europe, California, and South Africa. It was first described by Mathews (1971) from Northern Ireland. Known hosts are grasses (family Poaceae), principally Lolium perenne (perennial ryegrass), but also Dactylis glomerata (cat grass) and Festuca pratensis (meadow fescue) (Subbotin et al. 2010). Mowat (1974) reported that H. mani causes negligible damage to the yield of L. perenne in pot trials; however, Maas & Brinkman (1982) determined that it may cause significant damage to spring and autumn-sown perennial ryegrass in field conditions. During a routine examination for potato cyst nematode from a farm near Mawbanna in north-west Tasmania, Australia, several pale to dark brown Heterodera cysts were extracted that were lemon shaped with the presence of a small vulval cone at the posterior end and a distinct neck. The J2 (n=20) stylet length ranged from 24-26 µm with round knobs deeply concave anteriorly, hyaline tail length was 37-42 µm, true tail length ranged from 59-68 µm and total body length varied from 526-559 µm. All the above characters match those described for H. mani (Subbotin et al. 2010). To verify this identification, DNA was extracted from five individual J2 juveniles from a single cyst using QIAamp DNA micro kit (Qiagen®), and two gene regions amplified: internal transcribed spacer region of ribosomal RNA (ITS-rRNA) with primer pair AB28 and TW81 and cytochrome oxidase 1 (CO1) with primer pair JB3 and JB5 (Bowles et al. 1992; Curran et al. 1994; Derycke et al. 2005). One PCR reaction contained 10 µM (1 µl each) of each primer, 12.5 µl of OneTaq® DNA Polymerase and 5 µl of DNA template with a final volume of 25 µl. PCR products were sent for purification and Sanger sequencing at Macrogen (Seoul, Rep. of Korea). All resulting sequences were trimmed, aligned, and analysed using Geneious Prime® 2022.0.1 (www.geneious.com). Five ITS sequences (accessions ON402852-ON402856) and five CO1 sequences (accessions ON402857-ON402861) were submitted to GenBank. These ITS sequences were very similar to each other and exhibited 99.16-100% similarity with that of H. mani isolate from Hamminkeln, Germany (AY148377) (Subbotin et al. 2018). The CO1 sequences exhibited 98.96-100% similarity with that of H. mani isolate from Washington, USA (MG523097) (Subbotin et al. 2003). Obtained sequences were mapped to reference sequences downloaded from NCBI GenBank and maximum likelihood phylogenetic trees were calculated. Due to the lack of further living nematode material, pot experiments were not performed. Such experiments are not feasible in Tasmania currently and transfer of live nematode material to the Australian mainland presents logistic and legal issues. However, morphological and molecular evidence for species determination of H. mani was unequivocal and contributes to the list of cyst nematode species present in Australia. This is the first detection of H. mani in Australia and is a range extension of the species from North America, Africa, and Europe to Australia. The nematode may cause damage to perennial ryegrass in Australia, however, impact on yield still needs to be investigated.

The potato cyst nematode, Globodera rostochiensis, is a quarantine pest in Australia affecting a relatively small number of properties in three different production areas in the State of Victoria. The effects of susceptible (Trent, Sebago and Coliban) and H1-resistant potato cultivars (Atlantic, Crop 13 and Nicola) on nematode populations were compared in two trials in a naturally infested field in the 2008/09 season (Trial 1) and in the 2009/10 season (Trial 2). The latter included a bare fallow treatment. The reproduction factor Pf/Pi (final compared with initial nematode population) was used to determine treatment effects. Initial population density was very high, averaging 111 and 119 eggs g(-1) soil in Trial 1 and 2, respectively. The Pf/Pi of population density (eggs g(-1) soil) was greater after growing susceptible cultivars (average 1.74 and 2.92 in Trials 1 and 2, respectively) than after growing resistant cultivars (average 0.72 and 0.61 in Trials 1 and 2, respectively), or after a bare fallow (1.09) in Trial 2. This correlated with higher Pf/Pi values of cysts 500 g(-1) soil and eggs cyst(-1) for susceptible cultivars than for resistant cultivars. Average Pf/Pi values greater than one in both trials are consistent with more cysts and an increased population density after growing susceptible cultivars. There was a trend of population decline, that is, Pf/Pi < 1, after growing resistant cultivars (average Pf/Pi values of eggs cyst(-1) of 0.77 and 0.35 and of eggs g(-1) soil of 0.72 and 0.61, for Trials 1 and 2, respectively). However, Trial 2 showed that these Pf/Pi values were not significantly less than those for the bare fallow (0.68 eggs cyst(-1) and 1.09 eggs g(-1) soil). The susceptible cultivars Trent and Sebago produced lower yields than the resistant cultivars in both trials. In contrast, the susceptible Coliban yielded as well as the resistant cultivars, suggesting a high level of tolerance of this cultivar to infestation by the nematode. The resistant cultivar Crop 13 produced 34% and 36% greater total yields in Trials 1 and 2, respectively, than the once popular, but susceptible cultivar Sebago. This is the first report of the effects of potato cultivars on a population of the potato cyst nematode in an Australian field. The use of H1-resistant potato cultivars can feasibly reduce populations of G. rostochiensis Ro1 on infested land and reduce PCN risk on land regulated as "linked" with infested land.

Occurrence of cyst nematode Heterodera daverti Wouts and Sturhan 1978 (Wouts & Sturhan 1978) is reported from Australia for the first time. Cysts were recovered from soil samples collected from a hop farm in Merriang, Victoria. Morphological and morphometric characteristics of cysts and second stage juveniles match those described for this species reported from other parts of the world. Identification was further supported by molecular evidence through comparison of newly generated sequences of ITS rRNA, CO1 mtDNA and 28S rRNA gene regions with sequences previously available on NCBI GenBank.

The scope of this paper is limited to the taxonomy, detection, and reliable morphological and molecular identification of the potato cyst nematodes (PCN) Globodera pallida and G. rostochiensis. It describes the nomenclature, hosts, life cycle, pathotypes, and symptoms of the two species. It also provides detailed instructions for soil sampling and extraction of cysts from soil. The primary focus of the paper is the presentation of accurate and effective methods to identify the two principal PCN species.

Potato cyst nematodes (PCN) are damaging soilborne quarantine pests of potato in many parts of the world. There are two recognized species, Globodera pallida and G. rostochiensis, with only the latter species-the golden cyst nematode-present in Australia. PCN was first discovered in Australia in 1986 in Western Australia, where it was subsequently eradicated and area freedom for market access was reinstated. In Victoria, PCN was first detected in 1991 east of Melbourne. Since then, it has been found in a small number of localized regions to the south and east. Strict quarantine controls have been in place since each new detection. It has previously been speculated that there were multiple separate introductions of PCN into Victoria. Our study utilized a historic (years 2001 to 2014) PCN cyst reference collection to examine genetic variability of Victorian PCN populations to investigate potential historical origins and subsequent changes in the populations that might inform patterns of spread. DNA was extracted from single larvae dissected from eggs within cysts and screened using nine previously described polymorphic microsatellite markers in two multiplex polymerase chain reaction assays. Sequence variation of the internal transcribed spacer region of the DNA was also assessed and compared with previously published data. A hierarchical sampling strategy was used, comparing variability of larvae within cysts, within paddocks, and between local regions. This sampling revealed very little differentiation between Victorian populations, which share the same microsatellite allelic variation, with differences between local regions probably reflecting changes in allele frequencies over time. Our molecular assessment supports a probable single localized introduction into Victoria followed by limited spread to nearby areas. The Australian PCN examined appear genetically distinct from populations previously sampled worldwide; thus, any new exotic incursions, potentially bringing in additional PCN pathotypes, should be easily differentiated from existing established local PCN populations.

A species' diet and feeding strategy directly affect fitness and environmental interactions. Understanding spatial and temporal variation in diets can identify key resources, inform trophic relationships, and assist in managing threatened species. The nationally endangered Guthega skink, Liopholis guthega, is restricted to two isolated Australian alpine plateaux, the Bogong High Plains (BHP) in Victoria and Kosciuszko National Park (KNP) in New South Wales. We compared this species' foraging ecology over the summer period between these 'sky-islands' separated by similar to 100 km of lowland valleys. Scat composition did not differ between the two lizard populations, despite differences in the invertebrate assemblages present. However, L. guthega diet varied temporally over summer at both locations. Invertebrates, predominantly Hymenoptera and Coleoptera, were the dominant food group in early summer (78% volume (V), 100% frequency occurrence (F)) and mid-summer (80% V, 100% F). A significant dietary shift occurred in late summer, when lizards consumed predominantly plant material (63% V, 95.5% F), consisting primarily of seasonally abundant berries from the snow beard heath, Acrothamnus montanus. In contrast to similar-sized Egerniinae species, it appears L. guthega is capable of opportunistically shifting its diet towards plant material in response to temporal variation in resource availability. Furthermore, the prevalence of intact seeds in scats indicates L. guthega may play a significant role in seed dispersal. Understanding these trophic interactions will assist conservation management of L. guthega, allowing conditions for an already established captive colony to mimic the temporally variable diets present in situ, as well as informing revegetation initiatives aimed at maintaining and expanding wild populations.

Specimens of the Heath Sand-skipper butterfly Antipodia chaostola were found from only three locations in a survey in Central Gippsland in 1998. Th e preferred larval food plant, Thatch Saw-sedge Gahnia radula, is common and widespread but the butterfly appears restricted to areas of Heathy Woodland, which is a very uncommon vegetation community generally occurring on gentle, north-facing lower slopes. Th e dominant eucalypt species are stringybarks and peppermints, which commonly are stunted and sparse due to the very infertile, yellowish, gradational soils. Female Heath Sand-skippers appear to favour small young plants or regrowth of Saw-sedge, following bushfires, for oviposition. As fire may be important or necessary for new growth of Sawsedge, it consequently may be important or necessary for the persistence of the Heath Sand-skipper. Using a 9-12 year fuel reduction burn cycle, and staggering the burning of patches of the habitat in a mosaic pattern, seem necessary to provide a constant availability of appropriate habitat for the Heath Sand-skipper.

The Swamp Skink, Egernia coventryi, is an uncommon species that inhabits wetlands and swampy heaths in predominantly coastal regions of southeastern Australia. We examined museum specimens to quantify the diet, reproduction, and sexual dimorphism of E. coventryi. The mean SVL of both adult males and females was 85-86 mm, and individuals attain sexual maturity at about 72-74 mm SVL. Although the sexes did not differ in body size (SVL), males have longer and broader heads than females, both in terms of absolute head size and head size relative to body size. Females ovulate in September and October with parturition occurring in late January or early February. However, not all adult females collected during the breeding season were pregnant, suggesting that female E. coventryi may not always breed annually. Litter size in E. coventryi ranged from 1-4, with a mean litter size of 2.6. Egernia coventryti is omnivorous and a largely opportunistic forager, with spiders (found in 16 of 47 individuals, 34%), beetles (26%), lepidopterans (17%), ants (11%), hemipterans (9%) and aquatic amphipods (9%) the most common animal prey items found in the alimentary canals of individuals. However, plant materials (e.g., fruits/berries, seeds, other vegetation) also were found in the stomachs of the majority of the specimens (66%). Sloughed skin (26%) and the tail fragment of another E. coventryi were found in the alimentary tracts of preserved specimens. The majority of specimens (76%) were infested with endoparasites, with one specimen containing 130 nematodes, although the mean number of nematodes per specimen was 8.87.