Pathogenicity of indigenous entomopathogenic nematodes from Benin against mango fruit fly (Bactrocera dorsalis) under laboratory conditions

Bactrocera dorsalis fruit fly is the economically most significant tephritid pest species on Mango, Mangifera indica L., in Benin, and entomopathogenic nematodes (EPNs) represent good candidates for its control in the soil. In this study, the susceptibility of larvae and pupae of B. dorsalis to 12 EPN isolates originating from Benin was investigated. The effect of nematode concentrations (20, 50, 100, 200 and 300 Infective Juveniles (IJs)/B. dorsalis larva) and of different substrate moisture content (10, 15, 20, 25 and 30% v/w) on B. dorsalis mortality at the larval stage was studied. Also, the reproduction potential inside B. dorsalis larvae was assessed. Our results revealed that the susceptibility of B. dorsalis larvae was significantly different among the 12 tested nematode isolates with H. taysearae isolate Azohoue2 causing the greatest insect mortality (96.09 +/- 1.44%). The lowest insect mortality (7.03 +/- 4.43%) was recorded with Steinernema sp. strain Bembereke. Significant differences in insect mortality were recorded when EPNs were applied at varying IJs concentrations. A concentration of 100 nematodes of either H. taysearae Azohoue2 or H. taysearae Hessa1 per B. dorsalis larva was enough to kill at least 90% of B. dorsalis larvae. Larvae were less susceptible to nematodes at higher moisture content (25% and 30%). In addition, pupae were less susceptible to nematodes than larvae. Furthermore, the tested nematode isolates were able to reproduce inside B. dorsalis third instar larva with the Heterorhabditis isolates giving the greatest multiplication rate (59577.2 IJs +/- 14307.41)

Influence of pesticides, soil temperature and moisture on entomopathogenic nematodes from southern Benin and control of underground termite nest populations

The influence of three pesticides on the viability and infectivity of four Beninese isolates of entomopathogenic nematodes (EPN), Heterorhabditis indica Ayogbe1, H. sonorensis Azohoue2, H. sonorensis Ze3, and Steinernema sp. Bembereke, was determined. The impact of both soil temperature and soil moisture on the virulence of these EPN to Trinervitermes occidentalis was investigated in laboratory assays. The effect of EPN-infected Galleria mellonella larvae on underground populations of Macrotermes bellicosus was also examined. All tested Heterorhabditis species were more tolerant to glyphosate and fipronil than the Steinernema species. Heterorhabditis sonorensis Azohoue2, showed the best results with 63.2% termite mortality at a soil temperature of 35 degrees C. The increase of soil moisture to 20% (w/w) did not negatively influence the virulence of tested EPN. The underground populations of 71% or 60% treated nests were controlled by H. sonorensis Azohoue2-or H. indica Ayogbe1-infected G. mellonella larvae, respectively

Comparative susceptibility of Macrotermes bellicosus and Trinervitermes occidentalis (Isoptera: Termitidae) to entomopathogenic nematodes from Benin

The differential susceptibility of two termite species, Macrotermes bellicosus and Trinervitermes occidentalis, to four entomopathogenic nematodes (EPN) isolates from Benin, Heterorhabditis indica Ayogbel, H. sonorensis Azohoue2, H. sonorensis Ze3 and Steinernema sp. Bembereke, was bio-assayed in laboratory tests. Soldiers of both M. bellicosus and T occidentalis were similarly susceptible, but more susceptible than workers. Forty-eight h post-exposure of workers of M. bellicosus to 50 infective juveniles (IJ) of H. indica Ayogbel, H. sonorensis Azohoue2, H. sonorensis Ze3 and Steinernema sp. Bembereke for each termite resulted in 96.3, 87.9, 94.5 and 75.0% mortality, respectively, whereas under the same conditions, these EPN isolates caused 91.7, 98.5, 75.0 and 95.0% mortality of workers of T occidentalis. Soldiers of M. bellicosus were the most invaded with 13.2-18.6% of applied IJ. Based on concentration-mortality data, the isolates H. indica Ayogbel and H. sonorensis Ze3 were more virulent to M. bellicosus with LC50 values of 11 IJ, whereas Steinernema sp. Bembereke was the most virulent to T occidentalis with LC50 values of 12IJ. However, none of these isolates showed the highest penetration rate. All tested EPN isolates can recycle in both M. bellicosus and T occidentalis. Our EPN repellent-dispersing assay did not show evidence that M. bellicosus and T occidentalis would be able to detect the presence of D. of any EPN isolates/species. However, it was observed that nematode dispersal occurred by infected termites or phoresis

Molecular diversity of Photorhabdus and Xenorhabdus bacteria, symbionts of Heterorhabditis and Steinernema nematodes retrieved from soil in Benin

The diversity of 43 bacterial strains isolated from Beninese entomopathogenic nematodes was investigated molecularly by analyzing the 16S rRNA, recA, and gyrB genes. Based on 16S rRNA sequence analysis, 15 bacterial strains were identified as Xenorhabdus sp., 27 strains as Photorhabdus sp., and one as Serratia sp. The Xenorhabdus strains were isolated from Steinernema nematodes and identified as Xenorhabdus indica based on 16S rRNA gene and concatenated recA and gyrB sequence analysis. However, analysis of 16S rRNA and concatenated recA and gyrB gene sequences of the Photorhabdus strains, all isolated from Heterorhabditis nematodes, resulted in two separate sub-clusters (A) and (B) within the Photorhabdus luminescens group, distinct from the existing subspecies. They share low sequence similarities with nearest phylogenetic neighbors Photorhabdus luminescens subsp. luminescens Hb(T), Photorhabdus luminescens subsp. caribbeanensis HG29(T), and Photorhabdus luminescens subsp. noenieputensis AM7(T)

First record on the distribution of entomopathogenic nematodes (Rhabditida: Steinernematidae and Heterorhabditidae) in Southern Benin

For the first time, surveys of entomopathogenic nematodes (EPN) were conducted in five departments in the Guinean zone of Southern Benin. Out of 84 prospected sites and 280 collected soil samples from agricultural and natural vegetation, 26 (31.0%) and 32 (11.4%) were positive for EPN, respectively. Identification of the EPN was based on analyses of sequences of the ITS rDNA region and morphological/morphometric investigations. Two species were found, Heterorhabditis sonorensis and H. indica. This is the first record of H. sonorensis since its description from the Sonora desert in Mexico. Heterorhabditis sonorensis was the most common species, showing a preference for semi-closed habitats such as citrus orchards, other fruit production fields and woodland with soils having sand and organic matter content ranging between 53.6-89.5% and 0.1-4.7%, respectively, and a pH from acidic (4) to neutral (7.1). Entomopathogenic nematodes were not recovered from crop fields (maize, cassava, groundnut and bean) and soil samples with less than 50% sand content. Heterorhabditis indica was associated with citrus orchards and fruit fields on sand to sandy clay soils, with pH slightly acidic (pH = 5.4-6.4), but not with woodland. Discriminant analysis identified five major environmental variables, longitude, organic matter content and texture (silt, sand and clay content) to be the most important abiotic factors detemining the occurrence of EPN in soil from Southern Benin. Using these parameters, redundancy analysis revealed that H. sonorensis and H. indica prefer soils with high sand or organic matter content located in the more eastern longitude. No significant difference was observed in EPN species preferences taken individually, in terms of studied ecological parameters

Steinernema kandii n. sp. (Rhabditida: Steinernematidae), a new entomopathogenic nematode from northern Benin

Two nematode isolates from the genus Steinernema were collected in northern Benin. Morphological, morphometric, molecular and cross-hybridisation studies placed these nematodes into a new species, Steinernema kandii n. sp., within the bicornutum-group. Phylogenetic analyses based on both ITS and D2-D3 regions of 28S rDNA revealed that S. kandii n. sp. is different from all known Steinernema species and sister to S. abbasi (97.3-97.6% ITS nucleotide similarity) and S. bifurcatum (98.3-98.4% D2-D3 similarity). Steinernema kandii n. sp. can be separated from other members of the bicornutum-group by the greater infective juvenile (IJ) max. body diam. of 35 (27-48) mu m (type isolate). It differs from S. abbasi by the greater IJ body length 707 (632-833) mu m (type isolate), EP distance 55 (52-60) mu m (type isolate), spicule length 67 (57-75) mu m (type isolate) and the occurrence of one pair of genital papillae at the cloacal aperture

Evaluation of the ability of indigenous nematode isolates of Heterorhabditis taysearae and Steinernema kandii to control mango fruit fly Bactrocera dorsalis under laboratory, semi-field and field conditions in Northern Benin

We investigated the use of entomopathogenic nematodes to biologically control Bactrocera dorsalis in mango orchards. One isolate of Steinernema kandii (Thui) and two of Heterorhabditis taysearae (Hessa1 and Korobororou F4) were studied for their invasion time and virulence to third instar larvae of B. dorsalis in laboratory and semi field tests, respectively. In addition, the persistence of the same nematode isolates in soil under field conditions was tested. Results showed that all three nematode isolates could penetrate insect larvae after 2 h of exposure time. Furthermore, under semi field conditions, insect mortality was significantly different among EPN application times. The three nematode isolates were highly pathogenic to B. dorsalis with H. taysearae Hessa1 being the most virulent (70.84% +/- 10.46 [SEM] mortality) when EPNs were applied three days before insect introduction in the experimental pots. Moreover, Steinernema kandii persisted in soil up to 32 weeks after nematode application whereas both H. taysearae isolates persisted 30 weeks post application in the mango orchard. In general, four weeks upon nematode application, the density of infective juveniles decreased considerably and remained variable the following sampling dates