Peeyush Kumar

Repellent, larvicidal and pupicidal properties of essential oils and their formulations against the housefly, Musca domestica

The essential oils of six plant species [peppermint, Mentha piperita, and bergamot mint, Mentha citrata (both, Lamiales: Lamiaceae); blue gum, Eucalyptus globulus (Myrtales: Myrtaceae); lemongrass, Cymbopogon citratus, and khus grass, Vetiver zizanoides (both, Poales: Poaceae), and turmeric, Curcuma longa (Ziniberales: Zingiberaceae)] were screened for repellent, larvicidal and pupicidal activities against the housefly, Musca domestica L. (Diptera: Muscidae). Subsequently, emulsifiable concentrate (EC) formulations of the two most effective oils were prepared and tested in the laboratory as well as in the field. In repellency bioassays, M. piperita (RC84, 61.0 µg/cm2) was found to be most effective, followed by E. globulus (RC84, 214.5 µg/cm2) and C. citratus (RC84, 289.2 µg/cm2). Formulated M. piperita and E. globulus showed RC84 values of 1.6 µg/cm2 and 4.1 µg/cm2, respectively. Formulated M. piperita and E. globulus achieved larval mortality (LC50) in 72 h at 5.12 µg/cm2 and 6.09 µg/cm2, respectively. In pupicidal bioassays, crude oils of M. piperita and E. globulus suppressed the emergence of adult flies by 100%. Field experiments with the M. piperita formulation showed reductions in fly density (number of flies/h) of 96% on treated cattle and 98% on treated plots. This study demonstrates the effectiveness of EC formulations of selected essential oils in reducing housefly populations in field conditions.

Compositional analysis and insecticidal activity of Eucalyptus globulus (family: Myrtaceae) essential oil against housefly (Musca domestica).

The essential oil of Eucalyptus globulus (Myrtales: Myrtaceae) was evaluated for its chemical composition and insecticidal activity against the housefly, Musca domestica L. (Diptera: Muscidae). Chemical composition of E. globulus oil revealed 1,8-cineole (33.6%), α-pinene (14.2%) and d-limonene (10.1%) as major constituents, while vapour profile of E. globulus oil determined through solid phase microextraction (SPME) analysis showed 1,8-cineole (56.5%), α-pinene (16.9%), d-limonene (5.5%) and linalool acetate (3.4%) as principal components. Vapour phase of the oil showed increase in the contents of oxygenated monoterpenes. Insecticidal activity of E. globulus oil was assessed against larvae and pupae of housefly, through two different bioassays: contact toxicity and fumigation. Contact toxicity assay with larva showed lethal concentration, LC(50), between 2.73 and 0.60μl/cm(2) for different observation days while lethal time, LT(50), varied between 6.0 and 1.7 days. In fumigant assay for housefly larvae, LC(50) values of 66.1 and 50.1μl/l were obtained in 24h and 48h, respectively. Oil treated larvae showed surface shrinkage, spinous cells proliferation and bleb formation in scanning electron microscopy (SEM) observation. Pupicidal effectivity was measured in terms of percentage inhibition rate (PIR) which was 36.0-93.0% for contact toxicity and 67.9-100% for fumigation toxicity assay. Considerable activity of E. globulus oil against larvae and pupae of housefly demonstrates its potentiality as a viable option for the development of eco-friendly product for housefly control.

Biocontrol potential of essential oil monoterpenes against housefly, Musca domestica (Diptera: Muscidae).

Housefly (Musca domestica L.), one of the most common insects in human settlements, has been associated as vectors for various food-borne pathogens, causing food spoilage and disease transmission. The control of housefly was attempted using plant monoterpenes; menthone, menthol, menthyl acetate, limonene, citral and 1,8-cineole, against different life stages of housefly. Bioefficacy against housefly adults revealed highest repellent activity by menthol (95.6 percent) and menthone (83.3 percent). Against housefly larvae, menthol with an LC90 of 0.02 µl/cm(2) in contact toxicity assay and menthone with a LC90 value of 5.4 µl/L in fumigation assay were found to be most effective control agent. With respect to pupicidal activity, superior performance was shown by menthol, citral and 1,8-cineole in contact toxicity assay and citral and 1,8-cineole in fumigation assay. Limonene was found to be the poorest performer in all the assays. Overall, highest efficacy observed for menthol and menthone in various bioassays was in agreement with the results of essential oil activity obtained previously. Significant activity of monoterpenes against various life stages of housefly demonstrates their potential as excellent insecticides with prospects of monoterpenes being developed into eco-friendly and acceptable products for housefly control.

Preparation and characterization of PEG-Mentha oil nanoparticles for housefly control

Nanoparticles of Mentha × piperita essential oil were prepared by melt-dispersion method. The nanoparticles prepared at varying oil doses (5-10%, w/v) showed an encapsulation efficiency of 78.2-83.4%, while the oil load was observed to range between 3.64 and 7.46%. The average particle size of the nanoparticles varied between 226 and 331 nm, while polydispersity index showed variation between 0.547 and 1.000. DSC analysis indicated endothermic reaction during formation of nanoparticles, while a 2-term exponential kinetic model was followed during oil release. Nanoparticles showed considerable mortality against housefly larvae in lab (100%) as well as simulated field condition after first week (93%) and 6th week (57%) of application. This was the first study utilizing controlled release property of nanoparticles to formulate a cost effective product for breeding site application against housefly.

Comparative performance evaluation of multi-metal resistant fungal strains for simultaneous removal of multiple hazardous metals

In the present study, five fungal strains viz., Aspergillus terreus AML02, Paecilomyces fumosoroseus 4099, Beauveria bassiana 4580, Aspergillus terreus PD-17, Aspergillus fumigatus PD-18, were screened for simultaneous multimetal removal. Highest metal tolerance index for each individual metal viz., Cd, Cr, Cu, Ni, Pb and Zn (500mg/L) was recorded for A. fumigatus for the metals (Cd, 0.72; Cu, 0.72; Pb, 1.02; Zn, 0.94) followed by B. bassiana for the metals (Cd, 0.56; Cu, 0.14; Ni, 0.29; Zn, 0.85). Next, the strains were exposed to multiple metal mixture (Cd, Cr, Cu, Ni, Pb and Zn) of various concentrations (6, 12, 18, 30mg/L). Compared to other strains, B. bassiana and A. fumigatus had higher cube root growth (k) constants indicating their better adaptability to multi metal stress. After 72h, multimetal accumulation potential of B. bassiana (26.94±0.07mg/L) and A. fumigatus (27.59±0.09mg/L) were higher than the other strains at initial multimetal concentration of 30mg/L. However, considering the post treatment concentrations of individual metals in multimetal mixture (at all the tested concentrations), A. fumigatus demonstrated exceptional performance and could bring down the concentrations of Cd, Cu, Ni, Pb and Zn below the threshold level for irrigation prescribed by Food and Agriculture Organization (FAO).

Morpho-Molecular Characterization and Virulence Determination ofEntomopathogenic Fungal Isolates Native to Indian Subcontinent

Background: The house fly, Musca domestica L. is a major pest of medical and veterinary significance which due to their ubiquitous lifestyle and broad food preference achieves a very high density in favorable climatic conditions of Indian subcontinent. This has led to several instances of severe health condition among human and animal population, calling attention towards M. domestica control through existing and innovative methods. Use of biological control agent, such as entomopathogenic fungi (ENPF) for M. domestica control has shown potential, and need to be further explored for the newer isolates suited to particular environment for greater efficacy. The present study attempted the isolation and identification of native ENPF strains suitable for M. domestica control. Method and findings: Five strains of ENPF were isolated from soil samples, which on preliminary identification of morphological, microscopic and spore attributes were determined to be isolates of Beauveria species. The isolates showed wide variation in pathogenicity against M. domestica, with 72.7%-100% and 36.7%-72.3% mortality against adults and larvae life stage, respectively. Isolate ‘1’ depicting maximum insecticidal activity against M. domestica was selected for molecular identification using 18s-rRNA, and found to be Beauveria bassia HQ917687. The molecular analysis using random amplified polymorphic DNA (RAPD) showed isolate ‘2’ to be most similar to isolate ‘3’, while isolate ‘1’ and ‘5’ showed maximum variation between them. RAPD result was in conformity with various other properties of isolates; such as biomass, spore yield, hydrophobicity and insecticidal activity. Conclusion: The variations in activity obtained through pathogenicity assessment of fungal isolates in this study reflected the strain diversity of Beauveria isolates. The results also signified the importance of strain selection for effective control measures as well as for further consideration of commercial aspects.

Microscopic investigation to determine the effect of Beauveria bassiana (Bals.) Vuill. and Cymbopogon citratus (DC.) Stapf. treatment on different life stages of Musca domestica (L.)

Microscopic investigation was done to determine the effect of entomopathogenic fungi, Beauveria bassiana and essential oil of Cymbopogon citratus on different life stages of Musca domestica. Scanning electron microscopy investigation of fungal infected larvae showed sluggish movement, rigor, and failure of body to balance in water. Treated larvae also revealed varied level of cuticle shrinkage and extreme dehydration. Surface of B. bassiana infected pupae showed varied stage of mycelial growth, while the cadaver of adult fly was observed to have extensive fungal growth covering their entire body surface. The application of C. citratus oils on M. domestica larvae resulted in skin shrinkage, spinous cells proliferation and bleb formation, while the treated pupae showed high incidence of incomplete emergence and malformation in emerged adult flies. The current study establishes effect of C. citratus essential oil and B. bassiana infection on different life stages of M. domestica.

Evaluation of Beauveria bassiana infection in the hemolymph serum proteins of the housefly, Musca domestica L. (Diptera: Muscidae)

Beauveria bassiana plays a prominent role in biocontrol of houseflies, Musca domestica (L.). Thus, a deeper insight into immune response of M. domestica during B. bassiana infection was warranted to assist the production of more efficient mycoinsecticides. The present study investigates changes in protein profile of M. domestica hemolymph serum post B. bassiana infection using two-dimensional difference gel electrophoresis (2D-DIGE) followed by identification of selected proteins by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). The non-infected or control group of flies showed an expression of 54 proteins, while M. domestica infected with B. bassiana expressed a total of 68 hemolymph serum proteins. Thirty three proteins were expressed in both groups of houseflies, whereas 35 proteins were exclusively expressed in infected flies and 21 proteins were exclusively expressed in control flies. Among the 33 proteins which were expressed in both groups of houseflies, 17 proteins showed downregulation, while16 proteins were upregulated in the infected flies compared to the non-infected ones. The results from this study are expected to facilitate better understanding of insect’s immune response mechanism.