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Pathogenicity of Isolates of Serratia Marcescens towards Larvae of the Scarab Phyllophaga Blanchardi (Coleoptera).

Pineda-Castellanos ML, Rodríguez-Segura Z, Villalobos FJ, Hernández L, Lina L, Nuñez-Valdez ME - Pathogens (2015)

Bottom Line: Oral and injection bioassays using healthy Phyllophaga blanchardi larvae fed with the S. marcescens isolates showed different degrees of antifeeding effect and mortality.S. marcescens (Sm81) cell-free culture supernatant caused significant antifeeding effect and mortality to P. blanchardi larvae by oral bioassay and also mortality by injection bioassay.This insecticidal protein could have applications in agricultural biotechnology.

View Article: PubMed Central - PubMed

Affiliation: Centro de Investigación en Dinámica Celular, Instituto de Ciencias Básicas y Aplicadas, Universidad Autónoma del Estado de Morelos, Av. Universidad 1001, Col. Chamilpa, CP 62209, Cuernavaca, Morelos, Mexico. mlpc@uaem.mx.

ABSTRACT
Serratia marcescens is a Gram negative bacterium (Enterobacteriaceae) often associated with infection of insects. In order to find pathogenic bacteria with the potential to control scarab larvae, several bacterial strains were isolated from the hemocoel of diseased Phyllophaga spp (Coleoptera:Scarabaeidae) larvae collected from cornfields in Mexico. Five isolates were identified as Serratia marcescens by 16S rRNA gene sequencing and biochemical tests. Oral and injection bioassays using healthy Phyllophaga blanchardi larvae fed with the S. marcescens isolates showed different degrees of antifeeding effect and mortality. No insecticidal activity was observed for Spodoptera frugiperda larvae (Lepidoptera: Noctuidae) by oral inoculation. S. marcescens (Sm81) cell-free culture supernatant caused significant antifeeding effect and mortality to P. blanchardi larvae by oral bioassay and also mortality by injection bioassay. Heat treated culture broths lost the ability to cause disease symptoms, suggesting the involvement of proteins in the toxic activity. A protein of 50.2 kDa was purified from the cell-free broth and showed insecticidal activity by injection bioassay towards P. blanchardi. Analysis of the insecticidal protein by tandem- mass spectrometry (LC-MS/MS) showed similarity to a Serralysin-like protein from S. marcescens spp. This insecticidal protein could have applications in agricultural biotechnology.

No MeSH data available.


Related in: MedlinePlus

Injection bioassay showing insecticidal activity in the Sm81 and Sm65 cell-free culture supernatant and Sm81 50 kDa protein. Panel A. Supernatant from 24 h culture from the S. marcescens 81 and S. marcescens 65 (2 µg·mL−1) was injected into the larvae. Sm81H and Sm65H represent the same culture supernatant from Sm81 and Sm65 but boiled for 5 min. Sm81P represents the mortality observed for the larvae injected with the 50 kDa protein (2 µg·mL−1) purified from the cell-free culture supernatant from Sm81. The control represents a nutrient broth containing BSA (2 µg mL−1). Mortality was recorded for 22 days after injection. Differences among the groups of larvae shown by the letters above the lines were evaluated using χ2 (p < 0.0001; n = 14). Panel B. Dose-response bioassay showing dose dependency of the Sm81P50 insect killing activity. Dose-response curve for the injection bioassay of Sm81P50 (1 µg to 8 µg) against P. blanchardi larvae. Ten larvae were used for each dose.
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pathogens-04-00210-f004: Injection bioassay showing insecticidal activity in the Sm81 and Sm65 cell-free culture supernatant and Sm81 50 kDa protein. Panel A. Supernatant from 24 h culture from the S. marcescens 81 and S. marcescens 65 (2 µg·mL−1) was injected into the larvae. Sm81H and Sm65H represent the same culture supernatant from Sm81 and Sm65 but boiled for 5 min. Sm81P represents the mortality observed for the larvae injected with the 50 kDa protein (2 µg·mL−1) purified from the cell-free culture supernatant from Sm81. The control represents a nutrient broth containing BSA (2 µg mL−1). Mortality was recorded for 22 days after injection. Differences among the groups of larvae shown by the letters above the lines were evaluated using χ2 (p < 0.0001; n = 14). Panel B. Dose-response bioassay showing dose dependency of the Sm81P50 insect killing activity. Dose-response curve for the injection bioassay of Sm81P50 (1 µg to 8 µg) against P. blanchardi larvae. Ten larvae were used for each dose.

Mentions: To test the ability of the S. marcescens extracellular proteins to produce mortality by intracoelomic inoculation, a bioassay by injection was performed. Cell-free culture supernatant from the isolates Sm81 and Sm65 grown for 24 h was injected into healthy P. blanchardi larvae. As observed in Figure 4, control larvae injected with nutrient broth containing BSA cause no significant mortality. On the other hand, larvae injected with Sm81 and Sm65 cell-free culture supernatant showed a significant mortality of about 78% (p < 0.0001). No significant mortality was observed when the culture broth from both isolates was boiled and injected into the larvae.


Pathogenicity of Isolates of Serratia Marcescens towards Larvae of the Scarab Phyllophaga Blanchardi (Coleoptera).

Pineda-Castellanos ML, Rodríguez-Segura Z, Villalobos FJ, Hernández L, Lina L, Nuñez-Valdez ME - Pathogens (2015)

Injection bioassay showing insecticidal activity in the Sm81 and Sm65 cell-free culture supernatant and Sm81 50 kDa protein. Panel A. Supernatant from 24 h culture from the S. marcescens 81 and S. marcescens 65 (2 µg·mL−1) was injected into the larvae. Sm81H and Sm65H represent the same culture supernatant from Sm81 and Sm65 but boiled for 5 min. Sm81P represents the mortality observed for the larvae injected with the 50 kDa protein (2 µg·mL−1) purified from the cell-free culture supernatant from Sm81. The control represents a nutrient broth containing BSA (2 µg mL−1). Mortality was recorded for 22 days after injection. Differences among the groups of larvae shown by the letters above the lines were evaluated using χ2 (p < 0.0001; n = 14). Panel B. Dose-response bioassay showing dose dependency of the Sm81P50 insect killing activity. Dose-response curve for the injection bioassay of Sm81P50 (1 µg to 8 µg) against P. blanchardi larvae. Ten larvae were used for each dose.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4493471&req=5

pathogens-04-00210-f004: Injection bioassay showing insecticidal activity in the Sm81 and Sm65 cell-free culture supernatant and Sm81 50 kDa protein. Panel A. Supernatant from 24 h culture from the S. marcescens 81 and S. marcescens 65 (2 µg·mL−1) was injected into the larvae. Sm81H and Sm65H represent the same culture supernatant from Sm81 and Sm65 but boiled for 5 min. Sm81P represents the mortality observed for the larvae injected with the 50 kDa protein (2 µg·mL−1) purified from the cell-free culture supernatant from Sm81. The control represents a nutrient broth containing BSA (2 µg mL−1). Mortality was recorded for 22 days after injection. Differences among the groups of larvae shown by the letters above the lines were evaluated using χ2 (p < 0.0001; n = 14). Panel B. Dose-response bioassay showing dose dependency of the Sm81P50 insect killing activity. Dose-response curve for the injection bioassay of Sm81P50 (1 µg to 8 µg) against P. blanchardi larvae. Ten larvae were used for each dose.
Mentions: To test the ability of the S. marcescens extracellular proteins to produce mortality by intracoelomic inoculation, a bioassay by injection was performed. Cell-free culture supernatant from the isolates Sm81 and Sm65 grown for 24 h was injected into healthy P. blanchardi larvae. As observed in Figure 4, control larvae injected with nutrient broth containing BSA cause no significant mortality. On the other hand, larvae injected with Sm81 and Sm65 cell-free culture supernatant showed a significant mortality of about 78% (p < 0.0001). No significant mortality was observed when the culture broth from both isolates was boiled and injected into the larvae.

Bottom Line: Oral and injection bioassays using healthy Phyllophaga blanchardi larvae fed with the S. marcescens isolates showed different degrees of antifeeding effect and mortality.S. marcescens (Sm81) cell-free culture supernatant caused significant antifeeding effect and mortality to P. blanchardi larvae by oral bioassay and also mortality by injection bioassay.This insecticidal protein could have applications in agricultural biotechnology.

View Article: PubMed Central - PubMed

Affiliation: Centro de Investigación en Dinámica Celular, Instituto de Ciencias Básicas y Aplicadas, Universidad Autónoma del Estado de Morelos, Av. Universidad 1001, Col. Chamilpa, CP 62209, Cuernavaca, Morelos, Mexico. mlpc@uaem.mx.

ABSTRACT
Serratia marcescens is a Gram negative bacterium (Enterobacteriaceae) often associated with infection of insects. In order to find pathogenic bacteria with the potential to control scarab larvae, several bacterial strains were isolated from the hemocoel of diseased Phyllophaga spp (Coleoptera:Scarabaeidae) larvae collected from cornfields in Mexico. Five isolates were identified as Serratia marcescens by 16S rRNA gene sequencing and biochemical tests. Oral and injection bioassays using healthy Phyllophaga blanchardi larvae fed with the S. marcescens isolates showed different degrees of antifeeding effect and mortality. No insecticidal activity was observed for Spodoptera frugiperda larvae (Lepidoptera: Noctuidae) by oral inoculation. S. marcescens (Sm81) cell-free culture supernatant caused significant antifeeding effect and mortality to P. blanchardi larvae by oral bioassay and also mortality by injection bioassay. Heat treated culture broths lost the ability to cause disease symptoms, suggesting the involvement of proteins in the toxic activity. A protein of 50.2 kDa was purified from the cell-free broth and showed insecticidal activity by injection bioassay towards P. blanchardi. Analysis of the insecticidal protein by tandem- mass spectrometry (LC-MS/MS) showed similarity to a Serralysin-like protein from S. marcescens spp. This insecticidal protein could have applications in agricultural biotechnology.

No MeSH data available.


Related in: MedlinePlus