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Abstract. Crimean-Congo hemorrhagic fever (CCHF) is a vector-borne viral disease, widely distributed in different regions of the world. The fever is caused by the CCHF virus (CCHFV), which belongs to the Nairovirus genus and Bunyaviridae family. The virus is clustered in seven genotypes, which are Africa-1, Africa-2, Africa-3, Europe-1, Europe-2, Asia-1 and Asia-2. The virus is highly pathogenic in nature, easily transmissible and has a high case fatality rate of 10-40%. The reservoir and vector of CCHFV are the ticks of the Hyalomma genus. Therefore, the circulation of this virus depends upon the distribution of the ticks. The virus can be transmitted from tick to animal, animal to human and human to human. The major symptoms include headache, high fever, abdominal pain, myalgia, hypotension and flushed face. As the disease progresses, severe symptoms start appearing, which include petechiae, ecchymosis, epistaxis, bleeding gums and emesis. Enzyme-linked immunosorbent assay, quantitative polymerase chain reaction, antigen detection, serum neutralization and isolation of the virus by cell culture are the diagnostic techniques used for this viral infection. There is no specific antiviral therapy available thus far. However, ribavirin has been approved by the World Health Organization for the treatment of CCHFV infection. Awareness campaigns regarding the risk factors and control measures can aid in reducing the spread of this disease to a greater extent, particularly in developing countries.

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Chilo partellus is an invasive polyphagous pest that has not been effectively managed with chemical pesticides. To select potential dsRNAs for use in an alternate control strategy, it is crucial to identify and evaluate stable reference genes for knockdown expression studies. This study evaluates the expression stability of seven candidate reference genes in C. partellus larvae fed on crude bacterially-expressed dsRNAs and purified dsRNAs at different time intervals, as well as the developmental stages and sexes. The expression stabilities of the reference genes were evaluated with different software programmes, such as BestKeeper, NormFinder, deltaCt, geNorm, and RefFinder. The overall results rank ELF as the most stably expressed reference gene when larvae were fed with crude bacteria-induced dsRNAs and purified dsRNA. However, Tubulin and HSP70 were more stable under different developmental stages and sexes. The expression levels of larvae that were fed crude bacteria-induced dsRNAs of Chitinase and Acetylcholinesterase were normalized with the four most stable reference genes (ELF, HSP70, V-ATPase and Tubulin) and the least stable reference gene (18S and HSP70) based on the geNorm algorithm. The least stable reference gene showed inconsistent knockdown expression, thereby confirming that the validation of a suitable reference gene is crucial to improve assay accuracy for dsRNA-targeted gene selection in C. partellus.

Unnecessary weed growth in sugarcane fields forces plants to compete for nutrients and sunlight for survival, which most often leads to significant yield losses. As chemical herbicides cannot differentiate between crop plants and weeds, the development of herbicide-tolerant crops is anticipated. Four sugarcane varieties, CPF-234, CPF-213, HSF-240, and CPF-246, were used to develop glyphosate herbicide tolerance. A glyphosate-tolerant gene of 1368 bp cloned directionally under the 35S promoter with the β-glucuronidase (GUS) reporter gene was used as the transgene. Through the biolistic transformation of sugarcane, calli of all cultivars were transformed with glyphosate-tolerant gene constructs. Efficient regeneration conditions were optimized on 1.5-2.5 mg/L kinetin, 1.5-2.5 mg/L 6-benzylaminopurine (BAP), and 1 mg/L gibberellic acid (GA 3 ). The transformants exhibited the best shoot regeneration on a medium containing 2 mg/L kinetin, 2 mg/L BAP, 2 mg/L GA 3 , and 1 mg/L indole-3-acetic acid. Based on initial screenings through GUS assay, the transformation efficiency was 22%, 32%, 17%, and 13% for cultivars 246, 234, 213, and 240, respectively. In transformed sugarcane plants, the transgene of 1368 bp was amplified. ELISA with gene-specific coated monoclonal IgG confirmed the transgene protein expression. It was revealed that all acclimatized transgenic sugarcane plants survived the glyphosate spray application of 900 mL/0.404 ha, except for the control nontransformed plants. However, at spray application of 1100 mL/0.404 ha, transgenic plants having the transgene protein OD of 0.2 to 1.0 did not survive, while those that had a transgene protein OD range of between 1.0 and 2.0 did. In addition, weeds growing alongside transgenic sugarcane plants turned brown and subsequently died at glyphosate spray applications of both 900 and 1100 mL/0.404 ha.

Potato virus Y has emerged as a threatening problem in all potato growing areas around the globe. PVY reduces the yield and quality of potato cultivars. During the last 30 years, significant genetic changes in PVY strains have been observed with an increased incidence associated with crop damage. In the current study, computational approaches were applied to predict Potato derived miRNA targets in the PVY genome. The PVY genome is approximately 9 thousand nucleotides, which transcribes the following 6 genes:CI, NIa, NIb-Pro, HC-Pro, CP, and VPg. A total of 343 mature miRNAs were retrieved from the miRBase database and were examined for their target sequences in PVY genes using the minimum free energy (mfe), minimum folding energy, sequence complementarity and mRNA-miRNA hybridization approaches. The identified potato miRNAs against viral mRNA targets have antiviral activities, leading to translational inhibition by mRNA cleavage and/or mRNA blockage. We found 86 miRNAs targeting the PVY genome at 151 different sites. Moreover, only 36 miRNAs potentially targeted the PVY genome at 101 loci. The CI gene of the PVY genome was targeted by 32 miRNAs followed by the complementarity of 26, 19, 18, 16, and 13 miRNAs. Most importantly, we found 5 miRNAs (miR160a-5p, miR7997b, miR166c-3p, miR399h, and miR5303d) that could target the CI, NIa, NIb-Pro, HC-Pro, CP, and VPg genes of PVY. The predicted miRNAs can be used for the development of PVY-resistant potato crops in the future.