Hugues Baimey

Effectiveness of different Heterorhabditis isolates from Southern Benin for biocontrol of the subterranean termite, Macrotermes bellicosus (Isoptera: Macrotermitinae), in laboratory trials

The host-finding ability of 28 Heterorhabditis sonorensis isolates and one H. indica isolate from southern Benin and one H. indica isolate from India was examined in vertical migration sand bioassays against workers of a target citrus termite pest, Macrotermes bellicosus. Thereafter, nine selected isolates were subjected to further investigations on virulence. Our results revealed that both H. sonorensis and H. indica isolates exhibit a cruiser type of search strategy and were capable, to various degrees, of migrating, infecting and killing workers of M. bellicosus in sand columns up to 20 cm long over a period of 3 days. However, only three isolates of H. sonorensis caused 100% mortality to M. bellicosus at the greatest depth tested. The exotic H. indica isolate (LN2) did not show strong finding ability compared to the indigenous one. Concerning virulence, differences were observed among isolates for their ability to invade workers of M. bellicosus. After 12 h post exposure, H. sonorensis from Ze (Ze2) and H. sonorensis from Azohoue (Azohoue2) exhibited the lowest invasion time with IT50 = 3.35 and 3.67 h, respectively, and a higher penetration rate (11.4% and 10%, respectively) compared with the other isolates. In the concentration-mortality test, we found that, based on 95% confidence limits, all H. sonorensis and H. indica isolates appeared to be equal, with LC50 values ranging from nine to 16 infected juveniles (IJ) termite I. Interestingly, 40 II termite(-1) were enough to cause 80% mortality to M. bellicosus. Contrary to the LC50, the results of our studies clearly demonstrate that M. bellicosus exhibits a time-dependent susceptibility to the tested nematode isolates. So, the lowest LT50 was observed for H. sonorensis Ze2 (LT50 = 23.30 h), the highest for H. sonorensis Yokon (34.76 h). The LT50 of the indigenous H. indica isolate was estimated to 24.07 h. In addition, all selected isolates were able to reproduce in M. bellicosus workers. The highest reproduction potential in M. bellicosus was observed with H. sonorensis Yokon with 20 213 IJ/termite followed by H. sonorensis Ze2 with 19 368 IJ/termites. All tested Beninese EPN isolates were pathogenic to the citrus termite pest M. bellicosus, with H. sonorensis Ze2 being the most virulent.

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

Summary – The differential susceptibility of two termite species, Macrotermes bellicosus and Trinervitermes occidentalis, to four entomopathogenic nematodes (EPN) isolates from Benin, Heterorhabditis indica Ayogbe1, 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 Ayogbe1, 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 Ayogbe1 and H. sonorensis Ze3 were more virulent toM. bellicosus with LC50 values of 11 IJ, whereas Steinernema sp. Bembereke was the most virulent to T. occidentalis with LC50 values of 12 IJ. However, none of these isolates showed the highest penetration rate. All tested EPN isolates can recycle in both M. bellicosus and T. occidentalis .O ur EPN repellent-dispersing assay did not show evidence that M. bellicosus and T. occidentalis would be able to detect the presence of IJ of any EPN isolates/species. However, it was observed that nematode dispersal occurred by infected termites or phoresis.

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°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.

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 HbT, Photorhabdus luminescens subsp. caribbeanensis HG29T, and Photorhabdus luminescens subsp. noenieputensis AM7T.

Assessment of inoculation methods in evaluating response of yam cultivars to infection by Scutellonema bradys

Two glasshouse experiments were conducted to assess the influence of three inoculation methods on Scutellonema bradys multiplication on yam (Dioscorea spp.) and on growth and production of the crop. Three separate cultivars of yam were used in the study: two Dioscorea rotundata cultivars (Ala and Kpouna) and one Dioscorea cayenensis (Tabane). One-month-old plants were each inoculated with approximately 1000 S. bradys juveniles and adults. The inoculation methods included inoculation with chopped infected pieces of yam peel (about 0.5 × 0.3 cm2), chopped and blended infected yam peel and a water suspension (200 ml) of extracted nematodes compared with a S. bradys-free yam tuber peel control. The two experiments were harvested at 9 and 4 months after planting. Nematode population densities at harvest were significantly different (P ≤ 0.05) among inoculation methods and among yam cultivars. Higher nematode population densities were recorded on tubers from plants inoculated with unblended peel, followed by those inoculated with blended peel. Inoculation with nematode suspensions yielded the lowest nematode population densities. Inoculation with S. bradys did not affect the vine circumference and the number of tubers. Weights of tubers differed among inoculation methods for two of the three yam cultivars tested (P ≤ 0.05).

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) μm (type isolate). It differs from S. abbasi by the greater IJ body length 707 (632-833) μm (type isolate), EP distance 55 (52-60) μm (type isolate), spicule length 67 (57-75) μm (type isolate) and the occurrence of one pair of genital papillae at the cloacal aperture.