Pathogens and disease最新文献

Mycoplasma contamination of cell culture represents a serious problem in research and decontamination from cell-propagated obligate intracellular bacteria has proven challenging. Here, we presented an optimized protocol to remove Mycoplasma from contaminated Chlamydia trachomatis culture. A stepwise procedure of Mycoplasma removal entails (i) incubation in nonionic detergent-containing solution and (ii) separation of viable chlamydial organisms by fluorescence-activated cell sorting (FACS), followed by subcloning using a focus-forming assay. We also adapted a polymerase chain reaction (PCR) assay using paired universal and Mycoplasma-specific primers, which are distinguishable from the C. trachomatis counterparts, in combination with Sanger sequencing to determine the presence of mycoplasmas' 16S rRNA genes. These integrated approaches allow for full removal of Mycoplasma, as verified by the improved PCR assay, without compromising the capacity of viable C. trachomatis to adapt to new infection in epithelial cells. Some pitfalls during the Mycoplasma decontamination process are discussed.

Mycoplasma genitalium, the smallest prokaryotic microorganism capable of independent replication, is increasingly recognized as a sexually transmitted pathogen. M. genitalium protein of adhesion (MgPa) plays a pivotal role in the process of M. genitalium adhesion to host cells. We previously identified cyclophilin A as a cellular receptor of MgPa using the virus overlay protein binding assay (VOPBA) together with liquid chromatography-mass spectrometry (LC-MS). In the current study, we have evaluated H2B as an alternative cellular receptor for MgPa since H2B was assigned the second higher score as a potential binding partner of MgPa in the VOPBA and LC-MS screen. It was found that recombinant MgPa specifically bind to H2B both in the SV-HUC-1 cell membrane and in form of a recombinant protein. H2B was detected throughout the SV-HUC-1 cells, including the cytoplasmic membrane, cytosol and nucleus. Importantly, H2B partially inhibited the adhesion of M. genitalium to SV-HUC-1 cells. Finally, H2B was both co-precipitated with recombinant MgPa and co-localized with M. genitalium and recombinant MgPa in SV-HUC-1 cells. The above observations suggest that H2B may act as a potential cellular receptor of MgPa for mediating M. genitalium adhesion to host cells.

Interactions of Leishmania donovani secretory virulence factors with the host proteins and their interplay during the infection process in humans is poorly studied in Visceral Leishmaniasis. Lack of a holistic study of pathway level de-regulations caused due to these virulence factors leads to a poor understanding of the parasite strategies to subvert the host immune responses, secure its survival inside the host and further the spread of infection to the visceral organs. In this study, we propose a computational workflow to predict host-pathogen protein interactome of L.donovani secretory virulence factors with human proteins combining sequence-based Interolog mapping and structure-based Domain Interaction mapping techniques. We further employ graph theoretical approaches and shortest path methods to analyze the interactome. Our study deciphers the infection paths involving some unique and understudied disease-associated signaling pathways influencing the cellular phenotypic responses in the host. Our statistical analysis based in silico knockout study unveils for the first time UBC, 1433Z and HS90A mediator proteins as potential immunomodulatory candidates through which the virulence factors employ the infection paths. These identified pathways and novel mediator proteins can be effectively used as possible targets to control and modulate the infection process further aiding in the treatment of Visceral Leishmaniasis.

The fascinating discovery of the first giant virus, Acanthamoeba polyphaga mimivirus (APMV), belonging to the family Mimiviridae in 2008, and its associated virophage, Sputnik, have left the world of microbiology awestruck. To date, about 18 virophages have been isolated from different environmental sources. With their unique feature of resisting host cell infection and lysis by giant viruses, analogous to bacteriophage, they have been assigned under the family Lavidaviridae. Genome of T-27, icosahedral-shaped, non-enveloped virophages, consist of dsDNA encoding four proteins, namely, major capsid protein, minor capsid protein, ATPase and cysteine protease, which are essential in the formation and assembly of new virophage particles during replication. A few virophage genomes have been observed to contain additional sequences like PolB, ZnR and S3H. Another interesting characteristic of virophage is that Mimivirus lineage A is immune to infection by the Zamilon virophage through a phenomenon termed MIMIVIRE, resembling the CRISPR-Cas mechanism in bacteria. Based on the fact that giant viruses have been found in clinical samples of hospital-acquired pneumonia and rheumatoid arthritis patients, virophages have opened a novel era in the search for cures of various diseases. This article aims to study the prospective role of virophages in the future of human therapeutics.