Sapna Mishra

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.

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.

In vivo and in vitro control activity of plant essential oils against three strains of Aspergillus niger

Contamination of environment and food from the prevalent spores and mycotoxins of Aspergillus niger has led to several diseases in humans and other animals. The present study investigated the control activity of plant essential oils against three strains of A. niger. In the elaborate assays done through microdilution plate assay and agar disk diffusion assay in the lab condition and in vivo assay on the stored wheat grains, the essential oil of Thymus vulgaris depicted overall superior efficacy. In microdilution plate assay, the oil of Anethum graveolens showed best fungistatic activity, while best fungicidal activity was depicted by Syzygium aromaticum oil. The oil of T. vulgaris showed moderate control efficacy against A. niger strains with its antifungal activity resulting mainly due to killing of microorganism rather than growth inhibition. In agar disk diffusion assay, T. vulgaris oil with a zone of inhibition (ZOI) of 23.3–61.1% was the most effective fungicide. The in vivo assay to evaluate the protection efficacy of oils for stored wheat grains against A. niger (AN1) revealed T. vulgaris (90.5–100%) to be the best control agent, followed by the oil of S. aromaticum (61.9–100%). The GC–MS analysis of T. vulgaris oil indicated the presence of thymol (39.11%), γ-terpinene (19.73%), o-cymene (17.21%), and β-pinene (5.38%) as major oil components. Phytotoxic effects of the oils on wheat seeds showed no significant phytotoxic effect of oils in terms of seed germination or seedling growth. The results of the study demonstrated control potentiality of essential oils for the protection of stored wheat against A. niger with prospect for development of eco-friendly antifungal products.

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.

Development of Emulsified Concentrate (EC) formulation of Mentha piperita oil for control of mosquito larvae

Background Malaria is a leading cause of morbidity and mortality with an estimated 2.2 billion reported cases and 0.7 million annual deaths [1]. Continued research for malaria control although has resulted in significant reduction in malaria mortality rate, investigative measures are needed to address the escalating cost and threat of pesticide resistance. In this direction, biological control agent such as essential oil presents a good alternative [2-4]. However, the commercial applicability of essential oils for mosquitoes control is plagued by their slow action, variable efficacy and lack of suitable/user friendly end product [5]. Thus, in the present study Emulsified concentrate (EC) formulation of Mentha piperita oil, an effective mosquito larvicide [2,3], was prepared and characterized.