Myriam Elías-Santos

Effect of culture medium on the production and virulence of submerged spores of Metarhizium anisopliae and Beauveria bassiana against larvae and adults of Aedes aegypti (Diptera: Culicidae)

Three Metarhizium anisopliae and three Beauveria bassiana isolates were cultivated in media containing casamino acids, soybean flour or sunflower seed flour and were shaken for three days. M. anisopliae presented similar yields of around 106 submerged spores/ml without significant differences among them, whereas B. bassiana produced yields of around 108 spores/ml, of which GHA strain produced more submerged spores in the casamino acids medium. The other two strains showed no significant difference in the production of submerged spores in the three media used. Differences in mortality on Aedes aegypti larvae were observed with the submerged spores of Metarhizium depending on isolate and medium used. M. anisopliae 2157 caused significantly higher mortality (40%) when cultivated in casamino acids medium. It presented an LC50 of 8.93 × 105 submerged spores/ml water against mosquito larvae five days after application, whereas it caused 27% mortality in Ae. aegypti adults 10 days after application. In conclusion, fungal nutrition affected virulence of some isolates of M. anisopliae against Ae. aegypti larvae while such an effect was not noted for B. bassiana isolates.

Paecilomyces fumosoroseus blastospore production using liquid culture in a bioreactor

There are many advantages to using liquid cultures for the production of blastospores. These include mainly the processes of scale up which are relatively easy, as well as the control of parameters such as temperature, aeration and pH. In this work, we evaluated the production of Paecilomyces fumosoroseus blastospores using a low-cost liquid culture medium in a fermenter in comparison to a medium commonly used for this purpose, with regard to yield and viability of blastospores. The two media contained the same concentration of glucose but differed in N source (M1 containing casamino acids and M2 provided with collagen peptone and yeast extract). Starting with an inoculum of 1x106 blastospores/ml, M2 medium produced 2x1010 blastospores/ml after incubation for 72 h at 520 rev/min agitation and 1 v/v/m (volume air/volume liquid.min) aeration, while only 2.4 x 108/ml were produced with M1. In addition, the microorganisms in medium M1 grew more slowly during log phase and reached an earlier plateau at 36 h fermentation. The medium containing collagen peptone and yeast extract is an excellent alternative for the production of P. fumosoroseus blastospores, providing lower cost, higher yield and shorter propagation time, but formulation does need to be improved.

Effects of culture medium and formulation on the larvicidal activity of the mosquito pathogen Lagenidium giganteum (Oomycetes: Lagenidiales) against Aedes aegypti

In this work, we examined the production of infective zoospores of Lagenidium giganteum in four culture media, and the larvicidal activity of the cultures was determined against Aedes aegypti larvae, as well as the effect of polymer encapsulation. Medium containing sunflower seed extract showed the greatest production of zoospores, 5.92×10(6) zoospores/ml after six days of fermentation at 25±2°C and 150rpm shaking. This culture tested against A. aegypti 1st stage larvae caused different mortality rates at 24, 48 and 72h posttreatment. The LC(50) obtained was 43.9, 41.1 and 42.9μl of total culture/ml, at 24, 48 and 72h posttreatment respectively, while the culture grown in medium with soybean meal showed 3-5 times higher LC(50) values. Finally, the total culture including mycelium, zoospores and presporangia formulated with 2.5% pectin showed significantly higher mortality rates, around 100% more than the unformulated culture, whose values were from 40 to 1% at 3, 6, 9, and 12d posttreatment in the bioassays carried out in the laboratory to determine residual activity.

Evaluación de la Mortalidad Causada por Cepas Nativas de Bacillus thuringiensis y Formulados Preparados contra Trichoplusia ni, Spodoptera exigua, y Helicoverpa zea1

Resumen. Se evaluó la mortalidad causada por 24 aislamientos nativos de Bacillus thuringiensis Berliner provenientes de suelo de Yucatán contra los lepidópteros Trichoplusia ni (Hübner), Spodoptera exigua (Hübner), y Helicoverpa zea (Boddie). Los bioensayos preliminares contra larvas neonatas comparados con las cepas HD-1, GM-7, IB-7, e IB-62 permitieron seleccionaron 8 cepas nativas para la determinación de las CL50. En la comparación de medias de las CL50 se encontró a la cepa GM-IB-92 significativamente más tóxica contra T. ni y de toxicidad similar contra S. exigua en comparación con HD-1 y GM-7. Sin embargo, ninguna de las cepas nativas mostró toxicidad mayor a las cepas de referencia HD-1, GM-7, IB-7, e IB-62 contra H. zea. Dos formulaciones de la cepa GM-IB-92, preparadas con polímeros biodegradables y protectores de la luz ultra violeta fueron probadas y comparadas contra Dipel® 2X, donde presentó mayor actividad tóxica F2–5 contra T. ni. Las formulaciones preparadas mantuvieron su actividad tóxica por al menos 90 días de almacenamiento a 25–30°C. Los formulados preparados presentaron menor reducción en la actividad insecticida, del 4–13% que el extracto sin formular que mostró 54–88% después de la irradiación con luz uv por 72 horas.

Effect of Native Mexican Isolates of Isaria fumosorosea (Wize) Brown & Smith1 on Spodoptera exigua (Hübner) and Helicoverpa zea (Boddie)2

Abstract. The aim of this study was to evaluate pathogenicity and effect of four isolated native and a strain of Isaria fumosorosea on beet armyworm, Spodoptera exigua (Hübner), and corn earworm, Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae). Bioassays in a laboratory used neonate larvae of beet armyworm and corn earworm reared on artificial diet, which were inoculated by spraying or immersing with suspensions of 1 × 108 ml-1 conidia of I. fumosorosea, and subsequently incubated at 26°C, 65 ± 5% relative humidity, and 14:10 light: dark hours. Isolated HIB-30 killed the most immersed corn earworms, but fewest beet armyworms. HIB-30 most disrupted metamorphosis, 40%, by immersing beet armyworms, while HIB-29 and Pfr-612 least interrupted metamorphosis (8%), by spraying and immersing, respectively.

Evaluation of Heterorhabditis indica (Rhabditida: Heterorhabditidae) Nematode Strain from Sinaloa, Mexico, Against Bemisia tabaci Immatures Under Laboratory Conditions

Abstract. A native strain of Heterorhabditis indica Poinar, Karunakar & David nematode from Guasave, Sinaloa, Mexico, was identified by sequencing internal transcribed spacer regions of ribosomal DNA (rDNA). Assays against immature sweetpotato whitefly, Bemisia tabaci (Gennadius), used four strains of nematodes: Heterorhabditis bacteriophora Poinar Guatemala strain, H. bacteriophora Chiapas strain, a commercial product Steinernema feltiae (Filipjev) (Koppert®), and the H. indica native strain from Sinaloa. All strains at concentrations of 1,000, 750, 500, and 250 infective juveniles per milliliter, without significant differences among them, killed 70 to 95% of B. tabaci biotype B nymphs on leaves of tomato, Solanum lycopersicum (Mill), under laboratory conditions. This study is the first report of a native H. indica strain in Mexico, which is proposed as a biological control agent against B. tabaci.

Application of Isaria fumosorosea1 Blastospores Produced in Liquid Culture for Control of Bemisia argentifolii2 on Cotton Plants

Abstract. Blastospores of lsaria fumosorosea Wize grown in a bioreactor with casaminoacids or collagen peptone as the nitrogen source were evaluated for effectiveness against nymphs of silverleaf whitefly, Bemisia argentifolii Bellows & Perring, in cotton, Gossypium hirsutum L. The I. fumosorosea blastospores produced in the two liquid culture media were formulated with and without diatomaceous earth and sprayed onto terminal leaves of cotton in the field. The effectiveness of the materials was compared with Specifik®, a commercial formulation containing 1 × 1011 conidia per gram of I. fumosorosea formulated with Rhudo®, and a mixture of insecticides (Endosulfan® + Rescate® 20 PS). Blastospores produced in the medium containing casaminoacids as the main source of nitrogen and formulated with diatomaceous earth, as well as blastospores produced in the medium containing collagen peptone and not formulated with diatomaceous earth were most effective in reducing the number of nymphs. These experimental formulations killed 23–66% of the nymphs at 4 and 6 days posttreatment. Specifik® was less effective than the best experimental formulations and was effective only at 4 days post-application in the second experiment. The insecticides Endosulfan® plus Rescate® 20 PS used by some cotton growers were not effective on any sampling date.

Genetic variability analysis of entomopathogenic fungi isolated from citrus-growing areas of Mexico

The objective of the study was to isolate and characterize genetically native fungi that remain in the soil of the diverse citrus regions of Mexico. We collected 142 soil samples from different Mexican states: Campeche (13), Michoacan (15), Nuevo Leon (25), San Luis Potosi (25), Sinaloa (17), Sonora (17), Tabasco (5), Tamaulipas (20), and Yucatan (5). Larvae of Galleria mellonella were used as bait to detect, trap and multiply entomopathogenic fungi in vivo. 23% of the soil samples processed were positive for the presence of entomopathogenic fungi according to the macroscopic and microscopic characteristics: Beauveria was detected in 12% (17 isolates), Metarhizium in 1% (2 isolates) and Isaria in 10% (14 isolates). We analyzed the genetic variability of the fungi by using the sequences of internal transcribed ribosomal genes (ITS-5.8S-ITS2) and subsequently performed the digestion with the enzyme Hae III. The results of the PCR generated fragments of approximately 600 bp for Beauveria spp., and Metarhizium spp. In the case of Isaria spp., it amplified a fragment above of 600 bp. Both reference strains and field isolates produced similar restriction patterns. Genera of native entomopathogenic fungi were found on the diverse citrus areas of Mexico; this opens the field for the biological control of pests affecting citrus in each region.