Staphylococcus aureus is a common gram-positive human pathogen involved in both community-acquired and nosocomial infections ranging from localized superficial lesions to food poisoning and fatal systemic infections owing to its impressive array of virulence factors responsible for attaching, colonizing, invading, and avoiding host immune system. The discovery of antibiotics effectively checked the once deadly infections. However, resistance started soon after their discovery and first methicillin resistant strain of S. aureus was reported in early sixties. The most important attribute of MRSA resistance to penicllins is its acquisition of mecA gene coding for penicillin-binding protein PBP2a that blocks inhibitory action on peptidoglycan cross-linking. Now MRSA presents a serious global healthcare concern being responsible for prolonged hospital stays and increased mortality. The precise information of virulence factors and resistant traits of MRSA and their interplay in a community is key to minimize the intermixing of resistant and susceptible pathogens in the community

Klebsiella pneumoniae and Mycobacterium tuberculosis coinfection is one of the most lethal combinations that has been becoming frequent yet, not diagnosed and reported properly. Due to the simultaneous occurrence of both infections, diagnosis is delayed leading to inadequate treatments and mortality. With the rise of MDR Klebsiella and Mycobacterium, a prophylactic and an immunotherapeutic vaccine has to be entailed for preemptive and adroit therapeutic approach. In this study, we aim to implement reverse vaccinology approach that encompasses a comprehensive evaluation of vital aspects of the pathogens to explore immunogenic epitopes against Omp A of Klebsiella and Rv1698, Rv1973 of Mtb that may help in vaccine development. The designed multi-epitopic vaccine was assessed for antigenicity, allergenicity and various physiochemical parameters. Molecular docking and simulations were executed to assess the immunogenicity and complex stability of the vaccine. The final multi-epitopic vaccine is validated to be highly immunogenic and can serve as a valuable proactive remedy for subject pathogens.

Nosocomial infections caused by vancomycin-resistant Enterococcus have become a major problem. Bacteriophage therapy is proposed as a potential alternative therapy. Bacteriophages are viruses that infect bacteria and are ubiquitous in nature. Lytic bacteriophage was isolated from sewage water that infects VREF, the causative agent of endocarditis, bacteraemia, and urinary tract infections (UTIs). The phage produced clear plaques with unique clear morphology and well-defined boundaries. TEM results of phage revealed it to be 108 ± 0.2 nm long and 90 ± 0.5 nm wide. The characterization of bacteriophage revealed that infection process of phage was calcium and magnesium dependent and phage titers were highest under optimum conditions for VREF, with an optimal temperature range of 37–50°C. The maximum growth was observed at 37°C, hence having 100% viability. The latent period for phage was small with a burst size of 512 viral particles per bacterial cell. The phage was tested against various clinical strains and results proved it to be host specific. It can be used as a potential therapeutic agent for VREF infections. The phage efficiently eradicated VREF inoculated in cattle compost, poultry compost, and a soil sample which makes it a potential agent for clearing compost and soil sample.

Enterococcus faecium, is an important nosocomial pathogen with increased incidence of multidrug resistance (MDR) – specifically Vancomycin resistance. E. faecium constitutes the normal microbiota of the human intestine as well as exists in the hospitals and sewage, thus making the microorganism difficult to eliminate. Phage therapy has gained attention for controlling bacterial MDR infections and contaminations. We have successfully isolated from waste water and characterized a lytic bacteriophage STH1 capable of targeting Vancomycin resistant Enterococcus faecium (VREF) with high specificity. The phage was isolated from sewage water of a hospital at district Dera Ismail Khan, Pakistan. Initial characterization showed that magnesium and calcium ions significantly increased phage adsorption to the host. One step growth experiment showed a latent period of 18 min with burst size of 334 virions per cell. Optimal temperature and pH of the phage was 37°C and 7.0, respectively. Phage application to host strain grown in milk and water (treated and untreated) showed that the phage efficiently controlled bacterial growth. The study suggests that the phage STH1 can serve as potential control agent for E. faecium infections in medical facilities and in other environmental contaminations.

Acinetobacter baumannii is notorious for causing serious infections of the skin, lungs, soft tissues, bloodstream, and urinary tract. Despite the overwhelming information available so far, there has still been no approved vaccine in the market to prevent these infections. Therefore, this study focuses on developing a rational vaccine design using the technique of epitope mapping to curb the infections caused by A . baumannii . An outer membrane protein with immunogenic potential as well as all the properties of a good vaccine candidate was selected and used to calculate epitopes for selection on the basis of a low percentile rank, high binding scores, good immunological properties, and non-allergenicity. Thus, a 240 amino-acid vaccine sequence was obtained by manually joining all the epitopes in sequence-wise manner with the appropriate linkers, namely AAY, GPGPG, and EAAAK. Additionally, a 50S ribosomal protein L7/L12, agonist to the human innate immune receptors was attached to the N-terminus to increase the overall immune response towards the vaccine. As a result, enhanced overall protein stability, expression, immunostimulatory capabilities, and solubility of the designed construct were observed. Molecular dynamic simulations revealed the compactness and stability of the polypeptide construct. Moreover, molecular docking exhibited strong binding of the designed vaccine with TLR-4 and TLR-9. In-silico immune simulations indicated an immense increment in T-cell and B-cell populations. Bioinformatic tools also significantly assisted with optimizing codons which allowed for successful cloning of constructs into desired host vectors. Using in-silico tools to design a vaccine against A . baumannii demonstrated that this construct could pave the way for successfully combating infections caused by multidrug-resistant bacteria. Supplementary Information The online version contains supplementary material available at 10.1007/s10989-021-10316-7.