Dengue virus belonging to the family Flaviviridae and its four serotypes are responsible for dengue infections, which extend over 60 countries in tropical and subtropical areas of the world including Pakistan. During the ongoing dengue outbreak in Pakistan (2022), over 30,000 cases have been reported, and over 70 lives have been lost. The only commercialized vaccine against DENV, Dengvaxia, cannot be administered as a prophylactic measure to cure this infection due to various complications. Using machine learning and reverse vaccinology approaches, this study was designed to develop a tetravalent modified nucleotide mRNA vaccine using NS1, prM, and EIII sequences of dengue virus from Pakistani isolates. Based on high antigenicity, non-allergenicity, and toxicity profiling, B-cell epitope, cytotoxic T lymphocyte (CTL), and helper T lymphocyte (HTL) putative vaccine targets were predicted. Molecular docking confirmed favorable interactions between T-cell epitopes and their respective HLA alleles, while normal mode analysis validated high-affinity interactions of vaccine proteins with immune receptors. In silico immune simulations confirmed adequate immune responses to eliminate the antigen and generate memory. Codon optimization, physicochemical features, nucleotide modifications, and suitable vector availability further ensured better antigen expression and adaptive immune responses. We predict that this vaccine construct may prove to be a good vaccinal candidate against dengue virus in vitro as well.

Haemophilus influenzae is a Gram-negative bacterium characterized as a small, nonmotile, facultative anaerobic coccobacillus. It is a common cause of a variety of invasive and non-invasive infections. Among six serotypes (a–f), H. influenzae type b (Hib) is the most familiar and predominant mostly in children and immunocompromised individuals. Following Hib vaccination, infections due to other serotypes have increased in number, and currently, there is no suitable effective vaccine to induce cross-strain protective antibody responses. The current study was aimed to validate the capability of two 20-mer highly conserved synthetic tbp1 (transferrin-binding protein 1) peptide-based vaccine candidates (tbp1-E₁ and tbp1-E₂) predicted using in silico approaches to induce immune responses against H. influenzae strains. Cytokine induction ability, immune simulations, and molecular dynamics (MD) simulations were performed to confirm the candidacy of epitopic docked complexes. Synthetic peptide vaccine formulations in combination with two different adjuvants, BGs (Bacterial Ghosts) and CFA/IFA (complete/incomplete Freund’s adjuvant), were used in BALB/c mouse groups in three booster shots at two-week intervals. An indirect ELISA was performed to determine endpoint antibody titers using the Student’s t-distribution method. The results revealed that the synergistic use of both peptides in combination with BG adjuvants produced better results. Significant differences in absorbance values were observed in comparison to the rest of the peptide–adjuvant combinations. The findings of this study indicate that these tbp1 peptide-based vaccine candidates may present a preliminary set of peptides for the development of an effective cross-strain vaccine against H. influenzae in the future due to their highly conserved nature.

The fruit fly, Bactrocera zonata is found infesting more than 50 cultivated and wild fruit species and thus responsible for 25-100% of damages in fleshy fruits Pheromone and para-pheromone traps are used to examine the population extent of fruit flies in different locations of the world. Therefore, the present research was conducted to assess the synergistic effect of pheromones and citrus plant extracts. The experiment was performed in Y-tube olfactometer in laboratory conditions to observe the attraction/or repellence of B. zonata towards pheromones and citrus plant extracts separately and in combination. The study was conducted in Vector Ecology Laboratory, Department of Entomology, University of Agriculture, and Faisalabad. For odor preference, about six female peach fruit flies was released in a free choice wind tunnel containing host fruit. The relative attraction percentage toward each fruit was determined on the basis of total number of flies attracted to each host fruit and the process was repeated three times with one hour interval. To observe oviposition preferences about 20 male and female fruit flies adult of 14-21 days old and 24-48 hour starved was released in rearing cages (60 × 30 × 30 cm) for 24h with 250 g of each fruit. Host preference was evaluated on the basis of total number of pupae and emergence of adults from each fruit. The mean attraction and repellence behavior was analyzed using analysis of variance (ANOVA) and the Tuckey HSD test.

Chicken being white meat is superior to red meat for a number of reasons, such as its health benefits due to its low fat and cholesterol content. In the gastrointestinal tracts of chickens, there is a high concentration of complex microbial populations, mostly composed of bacteria but also includes fungus, viruses, bacterial, protozoa and microorganisms. The proposed study was conducted for Isolation and identification of bacteria from the intestine of chicken. For this purpose, ten samples of chicken’s intestine were collected and samples were divided into three experimental groups such as Control, T1 and T2. Each group contained ten specimens. The control group showed less numbers of colonies while T1 and T2 showed high colony count. Proper disinfection was maintained. During the whole trial, the bacterial colonies were counted from the both control and experimental groups T1 and T2 as control was outside while T1 and T2 were kept inside the incubator for 24 hours. T3 demonstrated the highest colony count as compared to T1 and control group. The minimum number of colonies observed on control group samples was 33 x 10 CFU/ml and maximum number of colonies were observed was 330 x 10 CFU/mL in control group. The number of colonies was observed in experimental group 1 was 180 x 10 CFU/mL and maximum number of colonies was 10 CFU/mL. In the experimental group 2 the minimum number 218 x 10 CFU/mL and maximum colonies were 510 x 10 CFU/mL. The data was obtained and subjected to statistical analysis by applying one-way ANOVA that showed highly significant differences as the p- value was found to be equal to 0.01 (p<0.05). Hence, after performing the following experiment, we isolated and identified the different bacterial species from intestine of Gallus gallus domesticus.

Dengue fever, a rapidly spreading mosquito-borne viral disease, poses a major global health issue, particularly in tropical and subtropical regions. The absence of a universally effective vaccine against all four dengue virus serotypes necessitates continued exploration of novel vaccine strategies. This study evaluated a PrM-derived multiple antigenic peptide (MAP) vaccine candidate in BALB/c mice. Molecular docking analyses predicted favourable interactions of the selected PrM-derived B-cell epitope with both human (human BCR Fab region PDB ID: 5IFH) and mouse (mouse BCR PDB ID: 8EMA) B-cell receptors, supporting its antigenic potential. ChemSketch was used to develop the PrM–MAP vaccine construct, which was then commercially synthesized. Mice received single, double, and triple booster doses of the vaccine, while control groups received either PBS or adjuvant alone. Humoral immune responses were measured by ELISA, and safety was evaluated through in vitro cytotoxicity assays on HEK293 cells and the in vivo histopathological examination of major organs. Mice immunized with a single booster dose produced significant levels of PrM-specific antibodies compared to subsequent booster doses, suggesting variability in the humoral response elicited. Optimal coating concentrations in ELISA demonstrated the importance of antigen concentrations in the sensitivity of the assay. The safety evaluation indicated high cell viability, up to 150 µg/mL of PrM–MAP, in HEK293 cells, and an absence of significant histopathological damage in treated animals. The PrM–MAP vaccine construct elicited a detectable humoral immune response while demonstrating a high safety profile in vitro and in vivo. These findings indicate the potential of MAP-based vaccine platforms as promising candidates for further development against the dengue virus.