Role of Protozoa in the Aerobic Purification of Sewage

DURING recent years, there has been increasing amount of evidence regarding the importance of protozoa in the aerobic purification of sewage1–4 and, more recently, some quantitative observations on protozoa in determining the condition of sludge and quality of effluent have been recorded5–10. The evidence so far obtained has, however, been only indirect, chiefly owing to the difficulty in separating the protozoa from the associated bacteria. This has now been achieved and the object of this note is to show that the isolated protozoa can bring about practically all the changes associated with the purification. The part played by the bacteria is almost negligible.

Location of Pathogenic Bacteria during Persistent Infections: Insights from an Analysis Using Game Theory

Bacterial persistent infections are responsible for a significant amount of the human morbidity and mortality. Unlike acute bacterial infections, it is very difficult to treat persistent bacterial infections (e.g. tuberculosis). Knowledge about the location of pathogenic bacteria during persistent infection will help to treat such conditions by designing novel drugs which can reach such locations. In this study, events of bacterial persistent infections were analyzed using game theory. A game was defined where the pathogen and the host are the two players with a conflict of interest. Criteria for the establishment of Nash equilibrium were calculated for this game. This theoretical model, which is very simple and heuristic, predicts that during persistent infections pathogenic bacteria stay in both intracellular and extracellular compartments of the host. The result of this study implies that a bacterium should be able to survive in both intracellular and extracellular compartments of the host in order to cause persistent infections. This explains why persistent infections are more often caused by intracellular pathogens like Mycobacterium and Salmonella. Moreover, this prediction is in consistence with the results of previous experimental studies.

Design of an HA2-based Escherichia coli expressed influenza immunogen that protects mice from pathogenic challenge

Influenza HA is the primary target of neutralizing antibodies during infection, and its sequence undergoes genetic drift and shift in response to immune pressure. The receptor binding HA1 subunit of HA shows much higher sequence variability relative to the metastable, fusion-active HA2 subunit, presumably because neutralizing antibodies are primarily targeted against the former in natural infection. We have designed an HA2-based immunogen using a protein minimization approach that incorporates designed mutations to destabilize the low pH conformation of HA2. The resulting construct (HA6) was expressed in Escherichia coli and refolded from inclusion bodies. Biophysical studies and mutational analysis of the protein indicate that it is folded into the desired neutral pH conformation competent to bind the broadly neutralizing HA2 directed monoclonal 12D1, not the low pH conformation observed in previous studies. HA6 was highly immunogenic in mice and the mice were protected against lethal challenge by the homologous A/HK/68 mouse-adapted virus. An HA6-like construct from another H3 strain (A/Phil/2/82) also protected mice against A/HK/68 challenge. Regions included in HA6 are highly conserved within a subtype and are fairly well conserved within a clade. Targeting the highly conserved HA2 subunit with a bacterially produced immunogen is a vaccine strategy that may aid in pandemic preparedness.

Prediction of protein-protein interactions between human host and a pathogen and its application to three pathogenic bacteria

Molecular understanding of disease processes can be accelerated if all interactions between the host and pathogen are known. The unavailability of experimental methods for large-scale detection of interactions across host and pathogen organisms hinders this process. Here we apply a simple method to predict protein-protein interactions across a host and pathogen organisms. We use homology detection approaches against the protein-protein interaction databases. DIP and iPfam in order to predict interacting proteins in a host-pathogen pair. In the present work, we first applied this approach to the test cases involving the pairs phage T4 - Escherichia coli and phage lambda - E. coli and show that previously known interactions could be recognized using our approach. We further apply this approach to predict interactions between human and three pathogens E. coli, Salmonella enterica typhimurium and Yersinia pestis. We identified several novel interactions involving proteins of host or pathogen that could be thought of as highly relevant to the disease process. Serendipitously, many interactions involve hypothetical proteins of yet unknown function. Hypothetical proteins are predicted from computational analysis of genome sequences with no laboratory analysis on their functions yet available. The predicted interactions involving such proteins could provide hints to their functions. (C) 2011 Elsevier B.V. All rights reserved.

A Coordinated Interdependent Protein Circuitry Stabilizes the Kinetochore Ensemble to Protect CENP-A in the Human Pathogenic Yeast Candida albicans

Unlike most eukaryotes, a kinetochore is fully assembled early in the cell cycle in budding yeasts Saccharomyces cerevisiae and Candida albicans. These kinetochores are clustered together throughout the cell cycle. Kinetochore assembly on point centromeres of S. cerevisiae is considered to be a step-wise process that initiates with binding of inner kinetochore proteins on specific centromere DNA sequence motifs. In contrast, kinetochore formation in C. albicans, that carries regional centromeres of 3-5 kb long, has been shown to be a sequence independent but an epigenetically regulated event. In this study, we investigated the process of kinetochore assembly/disassembly in C. albicans. Localization dependence of various kinetochore proteins studied by confocal microscopy and chromatin immunoprecipitation (ChIP) assays revealed that assembly of a kinetochore is a highly coordinated and interdependent event. Partial depletion of an essential kinetochore protein affects integrity of the kinetochore cluster. Further protein depletion results in complete collapse of the kinetochore architecture. In addition, GFP-tagged kinetochore proteins confirmed similar time-dependent disintegration upon gradual depletion of an outer kinetochore protein (Dam1). The loss of integrity of a kinetochore formed on centromeric chromatin was demonstrated by reduced binding of CENP-A and CENP-C at the centromeres. Most strikingly, Western blot analysis revealed that gradual depletion of any of these essential kinetochore proteins results in concomitant reduction in cellular protein levels of CENP-A. We further demonstrated that centromere bound CENP-A is protected from the proteosomal mediated degradation. Based on these results, we propose that a coordinated interdependent circuitry of several evolutionarily conserved essential kinetochore proteins ensures integrity of a kinetochore formed on the foundation of CENP-A containing centromeric chromatin.

Hydrolysis of aromatic beta-glucosides by non-pathogenic bacteria confers a chemical weapon against predators

Bacteria present in natural environments such as soil have evolved multiple strategies to escape predation. We report that natural isolates of Enterobacteriaceae that actively hydrolyze plant-derived aromatic beta-glucosides such as salicin, arbutin and esculin, are able to avoid predation by the bacteriovorous amoeba Dictyostelium discoideum and nematodes of multiple genera belonging to the family Rhabditidae. This advantage can be observed under laboratory culture conditions as well as in the soil environment. The aglycone moiety released by the hydrolysis of beta-glucosides is toxic to predators and acts via the dopaminergic receptor Dop-1 in the case of Caenorhabditis elegans. While soil isolates of nematodes belonging to the family Rhabditidae are repelled by the aglycone, laboratory strains and natural isolates of Caenorhabditis sp. are attracted to the compound, mediated by receptors that are independent of Dop-1, leading to their death. The b-glucosides-positive (Bgl(+)) bacteria that are otherwise non-pathogenic can obtain additional nutrients from the dead predators, thereby switching their role from prey to predator. This study also offers an evolutionary explanation for the retention by bacteria of `cryptic' or `silent' genetic systems such as the bgl operon.

Biogenic nanosilver incorporated reverse osmosis membrane for antibacterial and antifungal activities against selected pathogenic strains: An enhanced eco-friendly water disinfection approach

Reverse osmosis (RO) membranes have been used extensively in water desalination plants, waste water treatment in industries, agricultural farms and drinking water production applications. The objective of this work is to impart antibacterial and antifungal activities to commercially available RO membrane used in water purification systems by incorporating biogenic silver nanoparticles (AgNPs) synthesized using Rosa indica wichuriana hybrid leaf extract. The morphology and surface topography of uncoated and AgNPs-coated RO membrane were studied using Scanning Electron Microscopy (SEM) and Atomic Force Microscopy (AFM). Elemental composition of the AgNPs-coated RO membrane was analyzed by energy-dispersive X-ray spectroscopy (EDAX). The functional groups were identified by Fourier Transform Infrared spectroscopy (FT-IR). Hydrophilicity of the uncoated and AgNPs-coated RO membrane was analyzed using water contact angle measurements. The thermal properties were studied by thermogravimetric analysis (TGA). The AgNPs incorporated RO membrane exhibited good antibacterial and antifungal activities against pathogenic bacterial strains such as E. coli, S. aureus, M. luteus, K. pneumoniae, and P. aeruginosa and fungal strains such as Candida tropicalis, C. krusei, C. glabrata, and C. albicans.

Hybrid and Rogue Kinases Encoded in the Genomes of Model Eukaryotes

The highly modular nature of protein kinases generates diverse functional roles mediated by evolutionary events such as domain recombination, insertion and deletion of domains. Usually domain architecture of a kinase is related to the subfamily to which the kinase catalytic domain belongs. However outlier kinases with unusual domain architectures serve in the expansion of the functional space of the protein kinase family. For example, Src kinases are made-up of SH2 and SH3 domains in addition to the kinase catalytic domain. A kinase which lacks these two domains but retains sequence characteristics within the kinase catalytic domain is an outlier that is likely to have modes of regulation different from classical src kinases. This study defines two types of outlier kinases: hybrids and rogues depending on the nature of domain recombination. Hybrid kinases are those where the catalytic kinase domain belongs to a kinase subfamily but the domain architecture is typical of another kinase subfamily. Rogue kinases are those with kinase catalytic domain characteristic of a kinase subfamily but the domain architecture is typical of neither that subfamily nor any other kinase subfamily. This report provides a consolidated set of such hybrid and rogue kinases gleaned from six eukaryotic genomes-S. cerevisiae, D. melanogaster, C. elegans, M. musculus, T. rubripes and H. sapiens-and discusses their functions. The presence of such kinases necessitates a revisiting of the classification scheme of the protein kinase family using full length sequences apart from classical classification using solely the sequences of kinase catalytic domains. The study of these kinases provides a good insight in engineering signalling pathways for a desired output. Lastly, identification of hybrids and rogues in pathogenic protozoa such as P. falciparum sheds light on possible strategies in host-pathogen interactions.

Characterization of novel immunodominant antigens of Mycobacterium tuberculosis

Seven novel antigens of Mycobacterium tuberculosis, which had previously been identified based on reactivity to sera from patients with tuberculosis, were characterized. Nucleotide sequence analysis of the genes encoding these seven antigens identified one of them as the FtsH and a second as the aminoimidazole ribotide synthase of M. tuberculosis. Antisera raised to the recombinant forms of each of these seven antigens were used to study the distribution of these proteins within mycobacterial species as well as to determine their subcellular localization and hydrophobicity. Four of the seven antigens were conserved only among pathogenic strains of mycobacteria. Of the seven proteins studied, FtsH and a second protein of unknown identity were localized in membranes. Two were cytosolic, while two others, which had a high proline content, were tightly associated with the cell wall. One protein was secreted. This secreted protein could be identified by serum from a majority of tuberculosis patients but not BCG-vaccinated individuals, suggesting its potential use in the immunodiagnosis of tuberculosis.

Immunointervention for patients with HIV and tuberculosis

Therapeutic options aimed at confronting the HIV pandemic face many obstacles. Current opinion on HIV-induced pathogenic immune activation and strategies aimed at eliminating HIV, including a potential role for non-neutralising antibodies as part of a therapeutic vaccine option, was elegantly reviewed by Martin Cadogan and Angus Dalgleish. 1 It is important to note that, for eliciting such antibody responses in patients, functionally fit antigen presenting cells and effector T and B cells are cruc.

An enzymic approach to the study of sewage and sludges

The purpose of the work described here has been (a) to obtain some evidence on catalase (an oxidative enzyme) and protease, urease and phosphatase (hydrolytic enzymes) in sewage, activated sludge and septic tank sludge, and (b) to use this evidence, as a new approach, to find out the relationship between the main groups of the micro-organisms (bacteria and protozoa) and their relative influence on the purification process. To make a rapid assessment of the enzyme activities in these systems in the course of three weeks, as an experimental measure, rat tissues were added, which might serve as an additional or a ‘shock’ load of organic matter to follow broadly the development of bacteria and protozoa and the changes in the enzyme activities in the different systems. A control system with sewage alone was also run. The results showed that the initial decomposition of the fresh organic matter added to sewage and sludges was almost entirely due to bacterial activity and the later oxidative changes and removal of the suspended solids, including the bacteria, were largely due to the protozoa, such as Epistylis articulata. Analysis of the enzyme activities in the different materials showed, among other things, that the activated sludge, with its mized bacteria, protozoa and other organisms, as a whole, contained about twenty times more protease activity than an equivalent amount of the protozoan E. articulata, and that this protozoan contained five times more catalase activity than the activated sludge. The significance of these observations is discussed.

Studies on the Mechanism of Action of Miconazole: Effect of Miconazole on Respiration and Cell Permeability of Candida albicans

The antifungal drug, miconazole nitrate, inhibits the growth of several species of Candida. Candida albicans, one of the pathogenic species, was totally inhibited at a concentration of approximately 10 μg/ml. Endogenous respiration was unaffected by the drug at a concentration as high as 100 μg/ml, whereas exogenous respiration was markedly sensitive and inhibited to an extent of 85%. The permeability of the cell membrane was changed as evidenced by the leakage of 260-nm absorbing materials, amino acids, proteins, and inorganic cations. The results we present clearly show that the drug alters the cellular permeability, and thus the exogenous respiration becomes sensitive to the drug.