Two populations (CS19 and CS20) of entomopathogenic nematodes were isolated from the soils of veg... more Two populations (CS19 and CS20) of entomopathogenic nematodes were isolated from the soils of vegetable fields from Bijnor district, India. Based on morphological, morphometrical, and molecular studies, the nematodes were identified as Steinernema surkhetense. This work represents the first report of this species in India. The infective juveniles (IJs) showed morphometrical and morphological differences, with the original description based on longer IJs size. The IJs of the Indian isolates possess six ridges in their lateral field instead of eight reported in the original description. The analysis of ITS-rDNA sequences revealed nucleotide differences at 345, 608, and 920 positions in aligned data. No difference was observed in D2-D3 domain. The S. surkhetense COI gene was studied for the first time as well as the molecular characterization of their Xenorhabdus symbiont using the sequences of recA and gyrB genes revealing Xenorhabdus stockiae as its symbiont. These data, together with the finding of X. stockiae, suggest that this bacterium is widespread among South Asian nematodes from the ''carpocapsae'' group. Virulence of both isolates was tested on Spodoptera litura. The strain CS19 was capable to kill the larvae with 31.78 IJs at 72 hr, whereas CS20 needed 67.7 IJs.
Entomopathogenic nematodes can act as scavenger organisms and to investigate this phenomenon a ra... more Entomopathogenic nematodes can act as scavenger organisms and to investigate this phenomenon a range of possible hosts was exposed to Steinernema carpocapsae. In some cases, differences between an infected insect and a scavenged one were noticed. For example, when infective juveniles of S. carpocapsae were applied to arenas with live or dead locusts (Shistocerca gregaria), 97.5 % of the live ones were infected, and the nematodes reproduced throughout the body whereas only 19 % of the dead ones were scavenged and the reproducing nematodes were located only in the head of the insect cadavers. Other organisms such as Agriotes sp. were not affected by the nematodes when they were alive but around 28% were scavenged. With the differences found between infected and scavenged insects, it is proposed that some insects can be used as ecological markers for the study of scavenging in nature.
The life cycle of four Steinernema species was observed in 4 insect micro-insect host species (le... more The life cycle of four Steinernema species was observed in 4 insect micro-insect host species (less than 5mm long). Several parameters were measured: sex ratio of invading nematodes, percentages of host infection and offspring production, penetration rate of infective juveniles per insect and number of new generation of infective juveniles. All parameters varied among nematode species, micro-host species and application rates. All Steinernema species were capable to invade micro-insect hosts, however, invasion decreased as insect size decreased and as nematode species size increased. None of the nematode species achieved 100% mortality in the micro-hosts. Due to size differences in the nematode species, Steinernema glaseri was less capable of completing its life cycle and unable to invade smaller hosts whereas S. riobrave completed its life cycle in smaller hosts more frequently. The number of invading nematodes and the number of offspring produced had the same levels regardless of the nematode application rates, those results showed a maximum top in the number of individuals per micro-insect host. The offspring production in thrips species was only possible by endotokia matricida in S. riobrave. The sex of the invader nematodes also impeded the life cycle of S. affine because males colonized the entire body of the micro-insect host leaving no room for female invasion. The size of the host plays an irrefutable role in limiting the development of nematodes and it appears improbable that an entomopathogenic nematode population can persist in the soil without the presence of bigger insects.
The life cycle of entomopathogenic nematodes (EPN) occurs inside an insect cadaver and an accumul... more The life cycle of entomopathogenic nematodes (EPN) occurs inside an insect cadaver and an accumulation of ammonia initiates as a consequence of the nematodes defecation. This accumulation reduces the food resources quality and creates a detrimental environment for nematodes. When a given ammonia concentration is reached, the nematodes start their emergence process, searching for a new host. In the present work, this parameter, ammonia triggering point (ATP) was measured in 7 Steinernema species/strains. The effect of different ammonia concentrations on the recovery process and their consequences in the nematodes survival were also investigated. The results indicate that ATP varies among nematode species; Steinernema glaseri showed the highest ATP of the evaluated species (1.98±2.6 mg of NH4-N*g of Galleria mellonella(-1)); whereas Steinernema riobrave presented the lowest ATP (1.16±0.1 mg of NH4-N*g of G. mellonella(-1)). On the other hand, the nematode emergence could be a repulsive response when ATP is reached. As the ammonia concentration increased the recovery percentage of Steinernema feltiae (Chile strain) dropped gradually from 79.4±11.9% in the control treatment to 0% when 1mg of NH4-N*ml of bacterial broth(-1) was added. It is possible, that emergence process could be a repulsive response of the nematodes due to ammonia concentration when is reaching the ATP. The role of ammonia inside the insect cadavers, might suggests connections with some stages of the EPN life cycle.
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 2012
The use of Fourier transform mid-infrared spectroscopy with attenuated total reflection for chara... more The use of Fourier transform mid-infrared spectroscopy with attenuated total reflection for characterizing entomopathogenic bacteria from genera Xenorhabdus and Photorhabdus is evaluated for the first time. The resulting spectra of Xenorhabdus poinarii and Photorhabdus luminiscens were compared with the spectrum of Escherichia coli samples. The absorption spectra generated by the bacteria samples, were very different at the region below 1400 cm −1 which represents the stretching vibrations of phosphate and carbohydrates. Star diagrams of the fingerprint section of nematodes spectra (between 1350 and 1650 cm −1 ) for separation between spectra was used and showed to be a useful tool for classification purposes.
ABSTRACT Entomopathogenic nematodes have achieved a place in biological control programmes becaus... more ABSTRACT Entomopathogenic nematodes have achieved a place in biological control programmes because of their effectiveness, speed of action, innocuousness to non-insect targets and simplicity of mass production. However many challenges derived to the lack of knowledge in some critical steps from laboratories to their use in the fields, have to be resolved in order to improve their performance and to reduce the mass production costs. For those reasons, studies on entomopathogenic nematology have increased considerably in the last few decades. Also, there have been important changes in the ways that results are published; many of them relate to major transformations in scientific trends. Using bibliometric tools we characterize variations in number, types of journal, countries of origin, research topics and the number of participating countries, of 1923 papers (from 1980 to 2010) reported in several on-line editorial databases.
Archives Of Phytopathology And Plant Protection, 2012
ABSTRACT Frosty pod rot or moniliasis is a very destructive disease of cocoa plantations in Latin... more ABSTRACT Frosty pod rot or moniliasis is a very destructive disease of cocoa plantations in Latin America. The conventional methods to control this disease such as the use of chemical pesticides have failed or are too expensive for smallholders. Nowadays, biological alternatives are being investigated to improve yields and to keep moniliasis controlled. Xenorhabdus and Photorhabdus bacteria and their exudates were evaluated against Moniliophthora roreri. All bacteria demonstrated antifungal action reaching up to 97% after 13 days of exposure, whereas some exudates demonstrated more than 70%. Moniliophthora roreri did not sporulate in bacteria presence and sporulation was reduced by their exudates in 70%. The bacteria and exudates were compared with a commercial fungicide used by Venezuelan farmers; better performance of the bacteria and exudates was found and the fungicide was unable to control M. roreri. These experiments showed promising results of Xenorhabdus and Photorhabdus as biocontrol agents for being included in the integrated disease management programmes in Venezuela.
Morales-Montero, P., S. Flores, S. A. Subbotin and E. San-Blas. 2013. First report of Ditylenchus... more Morales-Montero, P., S. Flores, S. A. Subbotin and E. San-Blas. 2013. First report of Ditylenchus gallaeformans (Tylenchida: Anguinidae) inducing galls on Clidemia fendleri (Melastomataceae) from Venezuela. Nematropica 43:241-246.
Entomopathogenic nematodes cannot be considered only as parasitic organisms. With dead Galleria m... more Entomopathogenic nematodes cannot be considered only as parasitic organisms. With dead Galleria mellonella larvae, we demonstrated that these nematodes use scavenging as an alternative survival strategy. We consider scavenging as the ability of entomopathogenic nematodes to penetrate, develop and produce offspring in insects which have been killed by causes other than the nematode–bacteria complex. Six Steinernema and two Heterorhabditis species scavenged but there were differences among them in terms of frequency of colonisation and in the time after death of G. mellonella larvae that cadavers were penetrated. The extremes of this behaviour were represented by Steinernema glaseri which was able to colonise cadavers which had been freeze-killed 240 h earlier and Heterorhabditis indica which only colonised cadavers which had been killed up to 72 h earlier. Also, using an olfactometer, we demonstrated that entomopathogenic nematodes were attracted to G. mellonella cadavers.
Entomopathogenic nematodes complete their life cycles inside dead insects. The emergence of new i... more Entomopathogenic nematodes complete their life cycles inside dead insects. The emergence of new infective juveniles from the cadaver has been attributed (but never demonstrated) to food depletion or to the accumulation of metabolites from the breakdown of the host’s tissues. Here we give evidence that emergence is triggered by ammonia, a product of nematode defecation. We found that the emergence of Steinernemafeltiae infective juveniles from Galleriamellonella cadavers was stimulated by a particular level of ammonia. Emergence was delayed when ammonia in the cadaver was decreased and was prompted when increased. These findings will further improve the understanding of the nematode life cycle. Here we speculate that production of infective juveniles can be mediated by ammonia and work in a manner analogous to that of the dauer recovery inhibiting factor (DRIF) in Caenorhabditiselegans.
ABSTRACT Entomopathogenic nematodes are able to survive by scavenging. We tested Steinernema felt... more ABSTRACT Entomopathogenic nematodes are able to survive by scavenging. We tested Steinernema feltiae, S. affine and Heterorhabditis megidis alone or in different combinations to evaluate the responses of these nematodes when dead or live Galleria mellonella larvae were offered. Steinernema feltiae and S. affine scavenged upon dead G. mellonella larvae and about 30% more dead larvae were penetrated than live ones. By contrast, H. megidis penetrated more live larvae than dead ones. When the nematode species were combined, the results varied among the combinations, but the dead larvae were always used as a host. The behaviour of natural field populations of S. feltiae and S. affine was also compared. Steinernema feltiae showed no difference between scavenging and performing ‘normal infections’, whereas S. affine scavenged to a reduced amount (around 60% less); this difference could be related to the particular foraging strategy of these nematodes.
The use of Fourier transform mid-infrared spectroscopy with attenuated total reflection for chara... more The use of Fourier transform mid-infrared spectroscopy with attenuated total reflection for characterizing entomopathogenic nematodes is evaluated for the first time. The resulting spectra of Steinernema glaseri and Heterorhabditis indica were compared with the spectrum of Caenorhabditis elegans. In the absorption spectra generated by the nematodes samples, the absorption bands were assigned to the molecular species and some important components were identified including triglycerides, trehalose, glycogen and collagen. Also, the use of star diagrams for the fingerprint section of nematode spectra for separating genera is discussed.
Two populations (CS19 and CS20) of entomopathogenic nematodes were isolated from the soils of veg... more Two populations (CS19 and CS20) of entomopathogenic nematodes were isolated from the soils of vegetable fields from Bijnor district, India. Based on morphological, morphometrical, and molecular studies, the nematodes were identified as Steinernema surkhetense. This work represents the first report of this species in India. The infective juveniles (IJs) showed morphometrical and morphological differences, with the original description based on longer IJs size. The IJs of the Indian isolates possess six ridges in their lateral field instead of eight reported in the original description. The analysis of ITS-rDNA sequences revealed nucleotide differences at 345, 608, and 920 positions in aligned data. No difference was observed in D2-D3 domain. The S. surkhetense COI gene was studied for the first time as well as the molecular characterization of their Xenorhabdus symbiont using the sequences of recA and gyrB genes revealing Xenorhabdus stockiae as its symbiont. These data, together with the finding of X. stockiae, suggest that this bacterium is widespread among South Asian nematodes from the ''carpocapsae'' group. Virulence of both isolates was tested on Spodoptera litura. The strain CS19 was capable to kill the larvae with 31.78 IJs at 72 hr, whereas CS20 needed 67.7 IJs.
Entomopathogenic nematodes can act as scavenger organisms and to investigate this phenomenon a ra... more Entomopathogenic nematodes can act as scavenger organisms and to investigate this phenomenon a range of possible hosts was exposed to Steinernema carpocapsae. In some cases, differences between an infected insect and a scavenged one were noticed. For example, when infective juveniles of S. carpocapsae were applied to arenas with live or dead locusts (Shistocerca gregaria), 97.5 % of the live ones were infected, and the nematodes reproduced throughout the body whereas only 19 % of the dead ones were scavenged and the reproducing nematodes were located only in the head of the insect cadavers. Other organisms such as Agriotes sp. were not affected by the nematodes when they were alive but around 28% were scavenged. With the differences found between infected and scavenged insects, it is proposed that some insects can be used as ecological markers for the study of scavenging in nature.
The life cycle of four Steinernema species was observed in 4 insect micro-insect host species (le... more The life cycle of four Steinernema species was observed in 4 insect micro-insect host species (less than 5mm long). Several parameters were measured: sex ratio of invading nematodes, percentages of host infection and offspring production, penetration rate of infective juveniles per insect and number of new generation of infective juveniles. All parameters varied among nematode species, micro-host species and application rates. All Steinernema species were capable to invade micro-insect hosts, however, invasion decreased as insect size decreased and as nematode species size increased. None of the nematode species achieved 100% mortality in the micro-hosts. Due to size differences in the nematode species, Steinernema glaseri was less capable of completing its life cycle and unable to invade smaller hosts whereas S. riobrave completed its life cycle in smaller hosts more frequently. The number of invading nematodes and the number of offspring produced had the same levels regardless of the nematode application rates, those results showed a maximum top in the number of individuals per micro-insect host. The offspring production in thrips species was only possible by endotokia matricida in S. riobrave. The sex of the invader nematodes also impeded the life cycle of S. affine because males colonized the entire body of the micro-insect host leaving no room for female invasion. The size of the host plays an irrefutable role in limiting the development of nematodes and it appears improbable that an entomopathogenic nematode population can persist in the soil without the presence of bigger insects.
The life cycle of entomopathogenic nematodes (EPN) occurs inside an insect cadaver and an accumul... more The life cycle of entomopathogenic nematodes (EPN) occurs inside an insect cadaver and an accumulation of ammonia initiates as a consequence of the nematodes defecation. This accumulation reduces the food resources quality and creates a detrimental environment for nematodes. When a given ammonia concentration is reached, the nematodes start their emergence process, searching for a new host. In the present work, this parameter, ammonia triggering point (ATP) was measured in 7 Steinernema species/strains. The effect of different ammonia concentrations on the recovery process and their consequences in the nematodes survival were also investigated. The results indicate that ATP varies among nematode species; Steinernema glaseri showed the highest ATP of the evaluated species (1.98±2.6 mg of NH4-N*g of Galleria mellonella(-1)); whereas Steinernema riobrave presented the lowest ATP (1.16±0.1 mg of NH4-N*g of G. mellonella(-1)). On the other hand, the nematode emergence could be a repulsive response when ATP is reached. As the ammonia concentration increased the recovery percentage of Steinernema feltiae (Chile strain) dropped gradually from 79.4±11.9% in the control treatment to 0% when 1mg of NH4-N*ml of bacterial broth(-1) was added. It is possible, that emergence process could be a repulsive response of the nematodes due to ammonia concentration when is reaching the ATP. The role of ammonia inside the insect cadavers, might suggests connections with some stages of the EPN life cycle.
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 2012
The use of Fourier transform mid-infrared spectroscopy with attenuated total reflection for chara... more The use of Fourier transform mid-infrared spectroscopy with attenuated total reflection for characterizing entomopathogenic bacteria from genera Xenorhabdus and Photorhabdus is evaluated for the first time. The resulting spectra of Xenorhabdus poinarii and Photorhabdus luminiscens were compared with the spectrum of Escherichia coli samples. The absorption spectra generated by the bacteria samples, were very different at the region below 1400 cm −1 which represents the stretching vibrations of phosphate and carbohydrates. Star diagrams of the fingerprint section of nematodes spectra (between 1350 and 1650 cm −1 ) for separation between spectra was used and showed to be a useful tool for classification purposes.
ABSTRACT Entomopathogenic nematodes have achieved a place in biological control programmes becaus... more ABSTRACT Entomopathogenic nematodes have achieved a place in biological control programmes because of their effectiveness, speed of action, innocuousness to non-insect targets and simplicity of mass production. However many challenges derived to the lack of knowledge in some critical steps from laboratories to their use in the fields, have to be resolved in order to improve their performance and to reduce the mass production costs. For those reasons, studies on entomopathogenic nematology have increased considerably in the last few decades. Also, there have been important changes in the ways that results are published; many of them relate to major transformations in scientific trends. Using bibliometric tools we characterize variations in number, types of journal, countries of origin, research topics and the number of participating countries, of 1923 papers (from 1980 to 2010) reported in several on-line editorial databases.
Archives Of Phytopathology And Plant Protection, 2012
ABSTRACT Frosty pod rot or moniliasis is a very destructive disease of cocoa plantations in Latin... more ABSTRACT Frosty pod rot or moniliasis is a very destructive disease of cocoa plantations in Latin America. The conventional methods to control this disease such as the use of chemical pesticides have failed or are too expensive for smallholders. Nowadays, biological alternatives are being investigated to improve yields and to keep moniliasis controlled. Xenorhabdus and Photorhabdus bacteria and their exudates were evaluated against Moniliophthora roreri. All bacteria demonstrated antifungal action reaching up to 97% after 13 days of exposure, whereas some exudates demonstrated more than 70%. Moniliophthora roreri did not sporulate in bacteria presence and sporulation was reduced by their exudates in 70%. The bacteria and exudates were compared with a commercial fungicide used by Venezuelan farmers; better performance of the bacteria and exudates was found and the fungicide was unable to control M. roreri. These experiments showed promising results of Xenorhabdus and Photorhabdus as biocontrol agents for being included in the integrated disease management programmes in Venezuela.
Morales-Montero, P., S. Flores, S. A. Subbotin and E. San-Blas. 2013. First report of Ditylenchus... more Morales-Montero, P., S. Flores, S. A. Subbotin and E. San-Blas. 2013. First report of Ditylenchus gallaeformans (Tylenchida: Anguinidae) inducing galls on Clidemia fendleri (Melastomataceae) from Venezuela. Nematropica 43:241-246.
Entomopathogenic nematodes cannot be considered only as parasitic organisms. With dead Galleria m... more Entomopathogenic nematodes cannot be considered only as parasitic organisms. With dead Galleria mellonella larvae, we demonstrated that these nematodes use scavenging as an alternative survival strategy. We consider scavenging as the ability of entomopathogenic nematodes to penetrate, develop and produce offspring in insects which have been killed by causes other than the nematode–bacteria complex. Six Steinernema and two Heterorhabditis species scavenged but there were differences among them in terms of frequency of colonisation and in the time after death of G. mellonella larvae that cadavers were penetrated. The extremes of this behaviour were represented by Steinernema glaseri which was able to colonise cadavers which had been freeze-killed 240 h earlier and Heterorhabditis indica which only colonised cadavers which had been killed up to 72 h earlier. Also, using an olfactometer, we demonstrated that entomopathogenic nematodes were attracted to G. mellonella cadavers.
Entomopathogenic nematodes complete their life cycles inside dead insects. The emergence of new i... more Entomopathogenic nematodes complete their life cycles inside dead insects. The emergence of new infective juveniles from the cadaver has been attributed (but never demonstrated) to food depletion or to the accumulation of metabolites from the breakdown of the host’s tissues. Here we give evidence that emergence is triggered by ammonia, a product of nematode defecation. We found that the emergence of Steinernemafeltiae infective juveniles from Galleriamellonella cadavers was stimulated by a particular level of ammonia. Emergence was delayed when ammonia in the cadaver was decreased and was prompted when increased. These findings will further improve the understanding of the nematode life cycle. Here we speculate that production of infective juveniles can be mediated by ammonia and work in a manner analogous to that of the dauer recovery inhibiting factor (DRIF) in Caenorhabditiselegans.
ABSTRACT Entomopathogenic nematodes are able to survive by scavenging. We tested Steinernema felt... more ABSTRACT Entomopathogenic nematodes are able to survive by scavenging. We tested Steinernema feltiae, S. affine and Heterorhabditis megidis alone or in different combinations to evaluate the responses of these nematodes when dead or live Galleria mellonella larvae were offered. Steinernema feltiae and S. affine scavenged upon dead G. mellonella larvae and about 30% more dead larvae were penetrated than live ones. By contrast, H. megidis penetrated more live larvae than dead ones. When the nematode species were combined, the results varied among the combinations, but the dead larvae were always used as a host. The behaviour of natural field populations of S. feltiae and S. affine was also compared. Steinernema feltiae showed no difference between scavenging and performing ‘normal infections’, whereas S. affine scavenged to a reduced amount (around 60% less); this difference could be related to the particular foraging strategy of these nematodes.
The use of Fourier transform mid-infrared spectroscopy with attenuated total reflection for chara... more The use of Fourier transform mid-infrared spectroscopy with attenuated total reflection for characterizing entomopathogenic nematodes is evaluated for the first time. The resulting spectra of Steinernema glaseri and Heterorhabditis indica were compared with the spectrum of Caenorhabditis elegans. In the absorption spectra generated by the nematodes samples, the absorption bands were assigned to the molecular species and some important components were identified including triglycerides, trehalose, glycogen and collagen. Also, the use of star diagrams for the fingerprint section of nematode spectra for separating genera is discussed.
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Papers by Ernesto San-Blas