Quantification of pH tolerance levels among entomopathogenic nematodes.

Soil pH affects the availability of nutrients, which impacts plant growth and development. Similarly, soil pH may also influence microorganisms in the soil, either beneficial or nonbeneficial. One such group of beneficial microorganisms is entomopathogenic nematodes (EPN), parasites of soil-inhabiting insects. Entomopathogenic nematodes have a number of attributes that make them good alternatives to chemical insecticides. The objective of this study was to investigate pH tolerance of 11 steinernematids and six heterorhabditids post exposure to different pH levels. Entomopathogenic nematode populations were exposed to varying pH levels (pH2 to pH11) made up from two different chemical solutions (ammonium-acetate and citrate-phosphate). Entomopathogenic nematode populations are expected to have varying tolerance to different pH levels. The highest infective juvenile survival was obtained from pH3 to pH10 in citrate-phosphate, where all populations displayed >50% survival. Steinernema carpocapsae populations had >90% survival at pH3 to pH11 in citrate-phosphate solutions. Overall, the steinernematids had a higher survival range in ammonium-acetate pH solutions compared with the heterorhabditids. Moreover, Steinernema spp., S. carpocapsae (ScCxrd, ScAll, and ScItalian) and S. riobrave showed consistently higher survival in both acidic and alkaline solutions, when compared to the other steinernematids, suggesting that they may be applied in both acidic and alkaline soils. These findings can be of use when selecting EPNs for biological control purposes in the two countries, respectively.

Microbial control of phytophagous invertebrate pests in South Africa: Current status and future prospects.

Invertebrate pests pose a significant threat to food security on the African continent. In response, South Africa has become one of the largest importers of chemical pesticides in sub-Saharan Africa, with several hundred active ingredients registered. To address the over-reliance on such chemicals, the South African Department of Agriculture, Forestry and Fisheries (DAFF) has eliminated or restricted several pesticides since the late 1970s. The recent launch of the South African National Bio-Economy Strategy and establishment of the South African Bioproducts Organisation (SABO), together with new guidelines for registration of biopesticides in 2015, also support this endeavour. Concurrently, entomopathogen-related research and bioproduct development has increased over the past decade. Currently, 31 products (seven manufactured locally) are registered under the Fertilizers, Farm Feeds, Agricultural Remedies and Stock Remedies Act 36 of 1947. Commercially important microbes include Beauveria bassiana (Cordycipitaceae), Metarhizium anisopliae (Clavicipitaceae), Cydia pomonella granulovirus, Cryptophlebia leucotreta granulovirus, Helicoverpa armigera nucleopolyhedrovirus (Baculoviridae) and Bacillus thuringiensis subsp. kurstaki and B. thuringiensis subsp. aizawai (Bacillaceae). Both parasitic and entomopathogenic nematodes (EPNs) show potential for development as bioinsecticides with one commercial EPN product, based on Heterorhabditis bacteriophora (Heterorhabditidae), registered under the Act. Rapid scientific progression, supported by a favourable legislative environment, should facilitate further advances in microbial control of phytophagous invertebrate pests in South Africa.

Biological characterization of the entomopathogenic nematode, Steinernema innovationi: a South African isolate.

Entomopathogenic nematode species perform differently under different environmental conditions; therefore, the authors investigated the biological and environmental characteristics that could optimize performance of Steinernema innovationi. The authors studied the effect of temperature on infectivity and reproduction, the foraging behavior and host range. Thermal activity was optimal between 22 and 25°C. Highest infective juvenile (IJ) yields in last instar Galleria mellonella were observed at 22°C (333,014 IJs/g) and 25°C (354,165 IJs/g). An average of 26% of the IJs infected G. mellonella larvae at depths of 15 cm within 24 hr. Steinernema innovationi IJs raised greater than 95% of their body off the substrate and moved in a circular pattern, but did not jump. The hosts, Acheta domesticus, Chilo partellus, and Plutella xylostella showed the least susceptibility. All other hosts, [Eldana saccharina, Sesamia calamistis, Tenebrio molitor, G. mellonella, Cydia pomonella] suffered 100% larval mortality. Pupal mortality ranged from 47 to 68%. An LC50 and LC70 of 3 and 31 IJs/larva, respectively, were calculated for Agrotis ipsilon. The results indicate that S. innovationi is adapted to moderate temperatures. Furthermore, to the author's knowledge this is the first report of a nematode (particularly from 'glaseri group') that shows cruiser-type behavior in movement, but also nictates.Entomopathogenic nematode species perform differently under different environmental conditions; therefore, the authors investigated the biological and environmental characteristics that could optimize performance of Steinernema innovationi. The authors studied the effect of temperature on infectivity and reproduction, the foraging behavior and host range. Thermal activity was optimal between 22 and 25°C. Highest infective juvenile (IJ) yields in last instar Galleria mellonella were observed at 22°C (333,014 IJs/g) and 25°C (354,165 IJs/g). An average of 26% of the IJs infected G. mellonella larvae at depths of 15 cm within 24 hr. Steinernema innovationi IJs raised greater than 95% of their body off the substrate and moved in a circular pattern, but did not jump. The hosts, Acheta domesticus, Chilo partellus, and Plutella xylostella showed the least susceptibility. All other hosts, [Eldana saccharina, Sesamia calamistis, Tenebrio molitor, G. mellonella, Cydia pomonella] suffered 100% larval mortality. Pupal mortality ranged from 47 to 68%. An LC50 and LC70 of 3 and 31 IJs/larva, respectively, were calculated for Agrotis ipsilon. The results indicate that S. innovationi is adapted to moderate temperatures. Furthermore, to the author's knowledge this is the first report of a nematode (particularly from 'glaseri group') that shows cruiser-type behavior in movement, but also nictates.

Steinernema biddulphi n. sp., a New Entomopathogenic Nematode (Nematoda: Steinernematidae) from South Africa.

A new species of entomopathogenic nematode (EPN), Steinernema biddulphi n. sp., was isolated from a maize field in Senekal, Free State Province of South Africa. Morphological and molecular studies indicated the distinctness of S. biddulphi n. sp. from other Steinernema species. Steinernema biddulphi n. sp. is characterized IJs with average body length of 663 µm (606-778 µm), lateral fields with six ridges in mid-body region forming the formula 2,6,2. Excretory pore located anterior to mid-pharynx (D% = 46). Hyaline layer occupies approximately half of tail length. Male spicules slightly to moderately curved, with a sharp tip and golden brown in color. The first generation of males lacking a mucron on the tail tip while the second generation males with a short filamentous mucron. Genital papillae with 11 pairs and one unpaired preanal papilla. The new species is further characterized by sequences of the internal transcribed spacer (ITS) and partial 28S regions (D2-D3) of the ribosomal DNA (rDNA). Phylogenetic data show that S. biddulphi n. sp. belongs to the "bicornutum" clade within the Steinernematidae family.

Effect of Storage Temperature and Duration on Survival and Infectivity of Steinernema innovationi (Rhabditida: Steinernematidae).

Entomopathogenic nematode species differ in their optimum storage temperature; therefore, we conducted a study on the survival and infectivity of the recently described Steinernema innovationi from South Africa at five storage temperatures (5°C, 10°C, 15°C, 20°C, and 25°C) over 84 d using 20,000 infective juveniles (IJ) in 25 ml aqueous suspension containing 0.1% formalin. Our results showed that survival was highest and most stable at 15°C, ranging from 84% to 88% after 84 d. Infectivity of IJ against Galleria mellonella larvae was >90% for all temperatures except for 5°C at which survival decreased to 10% after 84 d. In addition, we stored 2.5 million IJ on a sponge formulation in 15 ml of 0.1% formalin solution for 84 d at the optimum 15°C followed by 2 wk storage at 25°C. Storage of the IJ on a sponge formulation for 14 d at 25°C post 15°C storage for 84 d did not have a detrimental effect on IJ survival (87%) or infectivity to G. mellonella (95%).

Steinernema tophus sp. n. (Nematoda: Steinernematidae), a new entomopathogenic nematode from South Africa.

A new entomopathogenic nematode, Steinernema tophus n. sp. is described from South Africa. Morphological, molecular (ribosomal gene sequence data) together with cross-hybridization studies were used for diagnostics and identification purposes. Both molecular and morphological data indicate the new species belongs to the 'glaseri-group' of Steinernema spp. Key morphological diagnostic traits for S. tophus n. sp. include the morphology of the spicules and gubernaculum. Morphometric traits of third-stage infective juveniles, including total body length (average 1,046 µm), tail length (average 70 µm), location of the excretory pore (average 92 µm), D% (average 63), E% (average 132) and H% (average 32) values are definitive. In addition to these morphological characters, analysis of rDNA (28S and ITS) gene sequences depict this Steinernema species as a distinct and unique entity.

Diversity and distribution of entomopathogenic nematodes (Steinernematidae, Heterorhabditidae) in South Africa.

A total of 1506 soil samples from different habitats in seven geographic regions of South Africa were evaluated for the presence of entomopathogenic nematodes (EPN). Nematodes were isolated from 5% of the samples. Among the steinernematids, four Steinernema sp. were recovered including Steinernema khoisanae and three new undescribed species. Although steinernematids were recovered from both humid subtropical and semiarid regions, this family accounted for 80% of EPN recovered from the semiarid climate zones characterised by sandy, acidic soils. Eight isolates of S. khoisanae were recovered from the Western Cape province. One of the new undescribed steinernematids (Steinernema sp. 1) was recovered only from the Free State and KwaZulu-Natal provinces where humid subtropical conditions prevail and soils are generally less acidic with higher clay content. A high level of adaptation, however, was noted with Steinernema sp. 2, which was recovered from a wide range of soil conditions and habitats ranging from semiarid (Western Cape province) to humid subtropical (KwaZulu-Natal province). A third undescribed steinernematid, Steinernema sp. 3, seemed better adapted to heavier soils with more than 80% of isolates recovered from fruit orchards in the Free State province. Heterorhabditis bacteriophora was the only heterorhabditid recovered during this survey. This species was particularly prevalent in four provinces ranging from humid subtropical to semiarid regions. Isolation of EPN directly from insect cadavers included Steinernema sp. 2 and one H. bacteriophora from an unidentified white grub (Scarabaeidae) cadaver (i.e., dual infection) and H. bacteriophora from the black vine weevil, Otiorhynchus sulcatus.