Cutaneous Leishmaniasis (CL), a neglected vector-borne disease caused by protozoan parasite Leishmania major (L. major), is a major public health concern, and the development of new strategies to reduce the disease incidence has become a top priority. Advances in immunoinformatics and in-silico epitope prediction could be a promising approach to designing a finest vaccine candidate. In this study, we aimed to design a peptide-based vaccine against CL using computational tools and identified ten B-cell-derived T-cell epitopes from the glycoprotein gp63 of L. major. All of the potential immunodominant epitopes were used to design a vaccine construct along with a linker and an adjuvant at the N-terminal for enhancing its immunogenicity. Additionally, many characteristics of the proposed vaccine were examined, and it was confirmed to be non-allergenic, non-toxic, and thermally stable. To assess the vaccine interaction with the innate immune toll-like receptor-4 (TLR-4), a 3D structure of the vaccine construct was developed. Molecular docking and molecular dynamic simulation were used to confirm the binding and to assess the stability of the vaccine-TLR4 complex and interactions, respectively. In conclusion, our multi-epitope vaccine will provide a gateway to analyze the protein function of a potential vaccine candidate against CL.

Scabies, a human skin infestation caused by the ectoparasitic mite Sarcoptes scabiei var. hominis, affects more than 200 million people globally. The prevailing knowledge of the disease process and host immune response mechanisms is limited. A better understanding of the host-parasite relationship is essential for the identification of novel vaccine and drug targets. Here we aimed to interrogate the transcriptomic profiles of mite-infested human skin biopsies with clinical manifestations of ordinary scabies subjects (“OS”; n = 05) and subjects naive to scabies (“control”; n = 03) using RNASeq data analysis. A combined clustering, network, and pathway mapping approach enabled us to identify key signaling events in the host immune and pro-inflammatory responses to S. scabiei infestation. The clustering patterns showed various differentially expressed genes including inflammatory responses and innate immunity genes (DEFB4A, IL-19, CXCL8, CSF3, SERPINB4, S100A7A, HRNR) and notably upregulation of the JAK-STAT pathway in scabies-infested samples. Mite-infested human skin biopsies (GSE178563) were compared with an ex-vivo porcine infested model (E-MTAB-6433) and human skin equivalents (GSE48459). Marked enrichment of immune response pathways (JAK-STAT signaling, IL-4 and IL-13 pathway, and Toll receptor cascade), chemokine ligands and receptors (CCL17, CCL18, CCL3L1, CCL3L3, CCR7), and cytokines (IL-13 and IL-20) were observed. Additionally, genes known for their role in psoriasis and atopic dermatitis were upregulated, e.g., IL-19. The detailed transcriptomic profile has provided an insight into molecular functions, biological processes, and immunological responses and increased our understanding about transcriptomic regulation of scabies in human.

Scabies is a human skin disease due to the burrowing ectoparasite Sarcoptes scabiei var. hominis resulting in intense itching and inflammation and manifesting as a skin allergy. Because of insufficient mite material and lack of in vitro propagation system for antigen preparation, scabies is a challenging disease to develop serological diagnostics. For allergen characterization, full-length S. scabiei tropomyosin (Sar s 10) was cloned, expressed in pET-15b, and assessed for reactivity with IgE antibodies from human sera. IgE binding was observed to Sar s 10 with sera collected from subjects with ordinary scabies, house dust mite (HDM)-positive and naive subjects and a diagnostic sensitivity of < 30% was observed. S. scabiei paramyosin (Sar s 11) was cloned, and expressed in pET-28a in three overlapping fragments designated Sspara1, Sspara2, and Sspara3. IgE and IgG binding was observed to Sspara2 and Sspara3 antigens with sera collected from ordinary scabies, and HDM-positive subjects, but no binding was observed with sera collected from naive subjects. Sspara2 displayed excellent diagnostic potential with 98% sensitivity and 90% specificity observed for IgE binding and 70% sensitivity for IgG. In contrast, the diagnostic sensitivity of Sspara3 was 84% for IgE binding and 40% for IgG binding. In combination, Sspara2 and Sspara3 provided an IgE sensitivity of 94%. This study shows that IgE binding to Sspara2 and Sspara3 is a highly sensitive method for diagnosis of scabies infestation in clinical practice. The developed enzyme-linked immunosorbent assay helps direct future development of a specific diagnostic tool for scabies.

Toxoplasmosis is a zoonotic disease caused by Toxoplasma gondii, a protozoan that infects warm-blooded animals and humans. Approximately one third of the global population is infected by T. gondii. We conducted a cross-sectional study to assess the risk factors and One Health knowledge of toxoplasmosis in Rawalpindi and Islamabad, Pakistan. From July through December 2020, we collected data using questionnaires. The results showed that 60% of participants had heard or read about the disease, 23.3% of participants had no knowledge about the disease, and 16.8% participants were not sure about the disease. More than half of the participants (53.3%) reported that toxoplasmosis was caused by toxins, 5.3% reported that toxoplasmosis was an animal disease, 13.8% reported that toxoplasmosis was a human disease, 65.8% reported that it was both an animal and human disease, and 15.3% reported that it was neither an animal nor a human disease. Approximately 80.5% of participants reported that individuals acquired toxoplasmosis by changing cat litter. Our study findings revealed a low level of knowledge and awareness about toxoplasmosis among males. Therefore, there should be awareness programs to educate individuals about the risks of this deadly disease and to provide information on the major routes of transmission.

Background: Plasmodium vivax contributes to over 70% malaria burden in Pakistan, but limited data exists on various aspects including genetic diversity of the parasite as compared to other parts of the world. Since the information about the genetic diversity of P. vivax assists to understand the population dynamics of the parasite, the current study was designed to understand population divergence of P. vivax in Pakistan using circumsporozoite protein (pvcsp) and merozoite surface protein-1 (pvmsp-1) genes as molecular markers. Methods: The PCR for pvcsp and pvmsp-1 genes was carried out for 150 P. vivax isolates, followed by DNA sequencing of 35 and 30, respectively. Genetic diversity and polymorphism were analysed using ChromasPro, ClustalW, MEGA7, DnaSP v.5 and WebLogo programs. Results: The PCR for pvcsp and pvmsp-1 genes was carried out for 150 P. vivax isolates and resulting the PCR products of 1100 bp for pvcsp and ~ 400 bp for pvmsp-1 genes, respectively. In the central-repeat region (CRR) of pvcsp gene, sequences comprised of four variable repeats of PRMs, out of which GDRADGQPA (PRM1), GDRAAGQPA (PRM2) were more extensively dispersed among the P. vivax isolates. Partial sequences (~ 400 bp) of block 2 of pvmsp-1 gene depicted high level of diversity. Conclusion: The results revealed the polymorphism and genetic diversity especially at the CRR of pvcsp and block 2 of pvmsp-1 genes, respectively. The base-line data presented here warrants future studies to investigate more into the genetic diversity of P. vivax with large sample size from across the country for better understanding of population dynamics of P. vivax that will help to control malaria at individual and community level.