Phospholipase C: underrated players in microbial infections.

During bacterial infections, one or more virulence factors are required to support the survival, growth, and colonization of the pathogen within the host, leading to the symptomatic characteristic of the disease. The outcome of bacterial infections is determined by several factors from both host as well as pathogen origin. Proteins and enzymes involved in cellular signaling are important players in determining the outcome of host-pathogen interactions. phospholipase C (PLCs) participate in cellular signaling and regulation by virtue of their ability to hydrolyze membrane phospholipids into di-acyl-glycerol (DAG) and inositol triphosphate (IP3), which further causes the activation of other signaling pathways involved in various processes, including immune response. A total of 13 PLC isoforms are known so far, differing in their structure, regulation, and tissue-specific distribution. Different PLC isoforms have been implicated in various diseases, including cancer and infectious diseases; however, their roles in infectious diseases are not clearly understood. Many studies have suggested the prominent roles of both host and pathogen-derived PLCs during infections. PLCs have also been shown to contribute towards disease pathogenesis and the onset of disease symptoms. In this review, we have discussed the contribution of PLCs as a determinant of the outcome of host-pathogen interaction and pathogenesis during bacterial infections of human importance.

Role of Streptococcus pneumoniae extracellular glycosidases in immune evasion.

Streptococcus pneumoniae (pneumococcus) typically colonizes the human upper airway asymptomatically but upon reaching other sites of the host body can cause an array of diseases such as pneumonia, bacteremia, otitis media, and meningitis. Be it colonization or progression to disease state, pneumococcus faces multiple challenges posed by host immunity ranging from complement mediated killing to inflammation driven recruitment of bactericidal cells for the containment of the pathogen. Pneumococcus has evolved several mechanisms to evade the host inflicted immune attack. The major pneumococcal virulence factor, the polysaccharide capsule helps protect the bacteria from complement mediated opsonophagocytic killing. Another important group of pneumococcal proteins which help bacteria to establish and thrive in the host environment is surface associated glycosidases. These enzymes can hydrolyze host glycans on glycoproteins, glycolipids, and glycosaminoglycans and consequently help bacteria acquire carbohydrates for growth. Many of these glycosidases directly or indirectly facilitate bacterial adherence and are known to modulate the function of host defense/immune proteins likely by removing glycans and thereby affecting their stability and/or function. Furthermore, these enzymes are known to contribute the formation of biofilms, the bacterial communities inherently resilient to antimicrobials and host immune attack. In this review, we summarize the role of these enzymes in host immune evasion.

Mycobacterial response to an acidic environment: protective mechanisms.

Given the emergence and spread of multidrug-resistant and extensively drug-resistant strains of Mycobacterium tuberculosis (Mtb), the world faces the urgency of finding new drugs to combat tuberculosis. Understanding the biochemical/physiological processes enabling Mtb to survive the stressful environment within macrophages and acquire tolerance, resistance and persistence against the stresses are the key to developing new approaches to tackle this health problem. As Mtb gains entry into the respiratory tract and is engulfed by macrophages, lowering pH acts as a primary defence of phagosomes within macrophages and also in the centres of caseating granulomas. It becomes essential for the pathogen to maintain pH homeostasis for survival in these conditions. Acid resistance mechanisms are well known and extensively studied in other bacteria such as Escherichia coli, Lactobacillus spp., Brucella spp., Helicobacter pylori and Listeria monocytogenes. However, in the case of Mtb, acid tolerance and resistance mechanisms still need to be explored in detail. This review aims to describe the current understanding of underlying mechanisms involved in countering low pH faced by Mtb as the acid resistance/tolerance mechanisms contribute to the pathogenesis of the disease.

Immunogenicity and cross-protective efficacy of recombinant H5HA1 protein of clade 2.3.2.1a highly pathogenic H5N1 avian influenza virus expressed in E.coli.

The global spread of H5N1 highly pathogenic avian influenza virus (HPAIV) in poultry has caused great economic loss to the poultry farmers and industry with significant pandemic threat. The current study involved production of recombinant HA1 protein of clade 2.3.2.1a H5N1 HPAIV (rH5HA1) in E.coli and evaluation of its protective efficacy in chickens. Purification under denaturing conditions and refolding by dialysis against buffers containing decreasing concentrations of urea was found to preserve the biological activity of the expressed recombinant protein as assessed by hemagglutination assay, Western blot and ELISA. The Montanide ISA 71 VGA adjuvanted rH5HA1 protein was used for immunization of chickens. Humoral response was maintained at a minimum of 4log2 hemagglutination inhibition (HI) titre till 154 days post 2nd booster. We evaluated the protective efficacy of rH5HA1 protein in immunized chickens by challenging them with homologous (2.3.2.1a) and heterologous (2.3.2.1c) clades of H5N1 HPAIV. In both the groups, the HI titre significantly increased (P < 0.05) after challenge and the virus shedding significantly (P < 0.05) reduced between 3rd and 14th day post challenge. The virus shedding ratio in oro-pharyngeal swabs did not differ significantly between both the groups except on 7 days post challenge and during the entire experimental period in cloacal swabs. These results indicate that rH5HA1 was able to induce homologous and cross protective immune response in chickens and could be a potential vaccine candidate used for combating the global spread of H5N1 HPAIV threat. To our knowledge, this is the first study to report immunogenicity and protective efficacy of prokaryotic recombinant H5HA1 protein in chicken.

Host phospholipase C-γ1 impairs phagocytosis and killing of mycobacteria by J774A.1 murine macrophages.

Macrophages represent the first line of defense against invading Mycobacterium tuberculosis (Mtb). In order to enhance intracellular survival, Mtb targets various components of the host signaling pathways to limit macrophage functions. The outcome of Mtb infection depends on various factors derived from both host and pathogen. A detailed understanding of such factors operating during interaction of the pathogen with the host is a prerequisite for designing new approaches for combating mycobacterial infections. This work analyzed the role of host phospholipase C-γ1 (PLC-γ1) in regulating mycobacterial uptake and killing by J774A.1 murine macrophages. Small interfering RNA mediated knockdown of PLC-γ1 increased internalization and reduced the intracellular survival of both Mtb and Mycobacterium smegmatis (MS) by macrophages. Down-regulation of the host PLC-γ1 was observed during the course of mycobacterial infection within these macrophages. Finally, Mtb infection also suppressed the expression of pro-inflammatory cytokine tumor necrosis factor-α and chemokine (C-C motif) ligand 5 (RANTES) which was restored by knocking down PLC-γ1 in J774A.1 cells. These observations suggest a role of host PLC-γ1 in the uptake and killing of mycobacteria by murine macrophages.