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Study on citronella essential oil (CEO) sensitivity of 217 microbial strains of 65 species, isolated from animals with different disease conditions, revealed that citronella oil inhibited growth of only 10.6% strains. CEO inhibited Candida but of no Aspergillus strain. CEO inhibited 22 of 211 bacterial strains. Ampicillin was the least effective antibiotic but inhibited 41.2% bacterial strains. Gram positive bacteria (GPBs) were 4.5 more sensitive (p, 0.0006) to CEO than Gram negative bacteria (GNBs). More GNB strains (p, 0.02) were multi-drug resistant (MDR) type than GPB strains. Probability of CEO resistant was high in MDR strains (p, 0.006). Most of the Brucella abortus strains had MDR (83.3%). Strains of swamp buffalo origin were more (p, 0.08) commonly CEO (96.6%) resistant than strains of dog (81.3%) origin. MDR was the maximum in abortion associated (51.2%) strains and minimum in diarrhoea associated strains (25%). The study indicated that CEO is not an effective antimicrobial against veterinary clinical isolates. Antimicrobial drug resistance and CEO resistance patterns of bacteria were dependent on type of pathogen, its source and association with disease in animals and may be important for deciding an effective antimicrobial therapy.

Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/ CRISPR associated protein (cas) are now being accepted as a highly specific method of gene editing. Among many other applications, CRISPR/cas has an immense potential to be used as antivirals. In this study, we successfully demonstrated CRISPR/Cas9 mediated inhibition of Bovine Herpes virus -1 (BHV-1) replication. BHV-1 causes economically important diseases in bovines with establishment of latency. Six essential genes and one non-essential gene of BHV-1 were targeted to assess the impact on virus replication. Inhibition of UL52, circ, and UL27 genes showed promising results, whereas the other three genes US6, UL18, and UL34 resulted in lower level of inhibition. Non-specific gene editing in host and virus was in-silico evaluated and was demonstrated by inhibition of virus induced apoptosis. Successful editing of one viral non-essential gene without any alterations in virus replication demonstrated the potential of CRISPR/Cas9 in replicating viral genome. Complete abrogation of virus replication was observed transiently (~24 hours post-transfection/hpt) when transfected with short lived in-vitro transcribed sgRNAs. Whereas, under constant expression of sgRNAs through plasmid, complete inhibition of virus replication was observed till ~72 hours post-infection. Complete inhibition of replication was also observed with in-vitro transcribed sgRNA when booster dose of sgRNA was trasnfected at 24hpt. It has been speculated that constant expression with plasmid based delivery may result in off-target activity which can be ruled out with short lived in-vitro transcribed sgRNA.

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