Involvement of Enterobacter cloacae in the mortality of the fish, Mugil cephalus

Aims:  To identify the causative agent of the mortality in the fish, Mugil cephalus , in Muttukadu lagoon.
Methods and Results:  An enteric bacterium from the kidneys of moribund fish M. cephalus , was isolated and identified as Enterobacter cloacae (MK). Mugil cephalus was experimentally infected by this isolate and was re-isolated from the kidneys of the moribund fish. Enterobacter cloacae isolates from the lagoon water (MW1, MW2 and reference strain ATCC 13047) and the reference strain were not able to induce similar pathogenesis. The putative factor imparting pathogenicity to the MK isolate was identified as a cationic molecule, which migrated towards the cathode on agarose gel electrophoresis.
Conclusions:  The Ent. cloacae (MK) isolate harbouring a cationic factor was the causative agent for the mortality of M. cephalus , found in Muttukadu lagoon.
Significance and Impact of the Study:  This study reveals that human enteric bacteria MK which is considered as nonpathogenic to fish, may become pathogenic to fish when it harbours this cationic factor. This cationic factor is found to be pathogenic to the fish M. cephalus leading to mortality. It was also found to be pathogenic to mice. Therefore, the shuttling of Ent. cloacae , harbouring cationic factor, between human and fish may be of human health importance.     

Structural basis for Rab11-mediated recruitment of FIP3 to recycling endosomes

The Rab11 GTPase regulates recycling of internalized plasma membrane receptors and is essential for completion of cytokinesis. A family of Rab11 interacting proteins (FIPs) that conserve a C-terminal Rab-binding domain (RBD) selectively recognize the active form of Rab11. Normal completion of cytokinesis requires a complex between Rab11 and FIP3. Here, we report the crystal structure and mutational analysis of a heterotetrameric complex between constitutively active Rab11 and a FIP3 construct that includes the RBD. Two Rab11 molecules bind to dyad symmetric sites at the C terminus of FIP3, which forms a non-canonical coiled-coiled dimer with a flared C terminus and hook region. The RBD overlaps with the coiled coil and extends through the C-terminal hook. Although FIP3 engages the switch and interswitch regions of Rab11, the mode of interaction differs significantly from that of other Rab-effector complexes. In particular, the switch II region undergoes a large structural rearrangement from an ordered but non-complementary active conformation to a remodeled conformation that facilitates the interaction with FIP3. Finally, we provide evidence that FIP3 can form homo-oligomers in cells, and that a critical determinant of Rab11 binding in vitro is necessary for FIP3 recruitment to recycling endosomes during cytokinesis.     

Phycobiliproteins as a commodity: trends in applied research,patents and commercialization

Phycobiliproteins are a group of colored proteins commonly present in cyanobacteria and red algae possessing a spectrum of applications. They are extensively commercialized for fluorescent applications in clinical and immunological analysis. They are also used as a colorant, and their therapeutic value has also been categorically demonstrated. However, a comprehensive knowledge and technological base for augmenting their commercial utilities is lacking. Hence, this work is focused towards this objective by means of analyzing global patents and commercial activities with application oriented research. Strategic mining of patents was performed from global patent databases resulting in the identification of 297 patents on phycobiliproteins. The majority of the patents are from USA, Japan and Europe. Patents are grouped into fluorescent applications, general applications and production aspects of phycobiliproteins and the features of each group are discussed. Commercial and applied research activities are compared in parallel. It revealed that US patents are mostly related to fluorescent applications while Japanese are on the production, purification and application for therapeutic and diagnostic purposes. Fluorescent applications are well represented in research, patents and commercial sectors. Biomedical properties documented in research and patents are not ventured commercially. Several novel applications are reported only in patents. The paper further pinpoints the plethora of techniques used for cell breakage and for extraction and purification of phycobiliproteins. The analysis identifies the lacuna and suggests means for improvements in the application and production of phycobiliproteins.     

Strengthening the laboratory diagnosis of pathogenic Corynebacterium species in the Vaccine era

Over the last three decades, successful implementation of the diphtheria vaccination in the developed and developing countries has reduced the infections caused by the toxigenic strains of Corynebacterium diphtheriae, but a concomitant increase in the invasive infections due to the nontoxigenic strains was seen. In addition, the recent reports on the emergence of nontoxigenic toxin gene‐bearing strains, having the potential to revert back to toxigenic form poses a significant threat to human beings. Besides infections caused by C. diphtheriae, the emergence of the respiratory, cutaneous and invasive infections by related pathogenic Corynebacterium species like C. ulcerans and C. pseudotuberculosis, complicate the diagnosis and management of infection. These observations together with the widespread prevalence of diphtheria in the vaccine era, necessitates the strengthening of the epidemiological surveillance and laboratory diagnosis of the pathogen. This review provides the overview of the advantages and limitations of different molecular methods and the role of MALDI‐TOF in the laboratory diagnosis of Diphtheria. The contribution of next generation sequencing technology and different genotyping techniques in understanding the pathogenicity, transmission dynamics and epidemiology of the C. diphtheriae is discussed.     

Development of low-cost media for the culture of mosquito larvicides, Bacillus sphaericus and Bacillus thuringiensis serovar. israelensis

Mosquito larvicides like Bacillus sphaericus and Bacillus thuringiensis serovar. israelensis have been widely and effectively used in mosquito control programs, but the industrial production of these bacilli is expensive. Here we have attempted to develop three cost-effective media, based on cheap sources, potato, common sugar and bengalgram. Growth and production of the insecticidal proteins from these bacteria were satisfactory. Bioassay studies with different mosquito larvae showed considerable toxicity. Therefore the investigation suggests that potato-based culture media are more economical for the industrial production of B. sphaericus and B. thuringiensis serovar. israelensis.     

All2: A tool for selecting mosaic mutations from comprehensive multi-cell comparisons

Accurate discovery of somatic mutations in a cell is a challenge that partially lays in immaturity of dedicated analytical approaches. Approaches comparing a cell’s genome to a control bulk sample miss common mutations, while approaches to find such mutations from bulk suffer from low sensitivity. We developed a tool, All², which enables accurate filtering of mutations in a cell without the need for data from bulk(s). It is based on pair-wise comparisons of all cells to each other where every call for base pair substitution and indel is classified as either a germline variant, mosaic mutation, or false positive. As All² allows for considering dropped-out regions, it is applicable to whole genome and exome analysis of cloned and amplified cells. By applying the approach to a variety of available data, we showed that its application reduces false positives, enables sensitive discovery of high frequency mutations, and is indispensable for conducting high resolution cell lineage tracing.     

SMS 2.0: an updated database to study the structural plasticity of short peptide fragments in non-redundant proteins

The function of a protein molecule is greatly influenced by its three-dimensional (3D) structure and therefore structure prediction will help identify its biological function. We have updated Sequence, Motif and Structure (SMS), the database of structurally rigid peptide fragments, by combining amino acid sequences and the corre-sponding 3D atomic coordinates of non-redundant (25%) and redundant (90%) protein chains available in the Protein Data Bank (PDB). SMS 2.0 provides information pertaining to the peptide fragments of length 5-14 resi-dues. The entire dataset is divided into three categories, namely, same sequence motifs having similar, intermedi-ate or dissimilar 3D structures. Further, options are provided to facilitate structural superposition using the pro-gram structural alignment of multiple proteins (STAMP) and the popular JAVA plug-in (Jmol) is deployed for visualization. In addition, functionalities are provided to search for the occurrences of the sequence motifs in other structural and sequence databases like PDB, Genome Database (GDB), Protein Information Resource (PIR) and Swiss-Prot. The updated database along with the search engine is available over the World Wide Web through the following URL http://cluster.physics.iisc.ernet.in/sms/.     

Immobilization of the fungus, Tolypocladium sp. for the production of Cyclosporin A

The fungus Tolypocladium sp. obtained from the culture collection of VCRC, Pondicherry, was immobilized in calcium alginate and used for the production of Cyclosporin A in a packed bed reactor under batch and continuous flow modes. The precursor amino acids, L-valine and L-leucine increased the yield of Cyc A when added individually; however when added together, the increase in the yield was no better than as added individually. The half-life of the biocatalyst was found to be around six months.     

Differential sensitivity of cytotoxic T lymphocytes and lymphokine-activated killer cells to inhibition by L-ornithine

The selective inhibition of murine cytotoxic T lymphocyte (CTL) differentiation in C57B1/6 (B6) anti-DBA/2 mixed leukocyte cultures (MLC) by the amino acid L-ornithine (Orn) could not be reversed by addition of up to 1000 U/ml IL-2. Analysis of the effects of Orn on induction of lymphokine-activated killer (LAK cells), using dosages of IL-2 from 10-1000 U/ml and measuring cytolytic activity against two tumor targets (P815 and YAC-1) over the course of 5 days, indicated that LAK cells were not suppressed by Orn. LAK precursors and effector cells were CD8- and ASGM1+, indicating that they were derived from natural killer (NK) cells. We also found that the growth and maintenance of cloned CTL lines were not sensitive to inhibition by Orn; nor was their acquisition of nonspecific cytolytic activity in the presence of high lymphokine concentrations. Thus, induction of naive CTL shows differential susceptibility to Orn inhibition relative to LAK and LAK-like activities by NK and cloned CTL lines in response to IL-2.     

Induction of Bacillus subtilis sporulation by nucleosides: inosine appears to be sporogen.

Inosine completely reversed the selective inhibition of sporulation in Bacillus subtilis 168 caused bym-aminobenzeneboronic acid; guanosine and adenosine, but not xanthosine, partially reversed inhibition, whereas pyrimidine nucleosides were slightly effective. In addition, 0.005 to 0.025 mM inosine caused a four- to fivefold stimulation of sporulation of B. subtilis grown in minimal salts medium. Ultraviolet and infrared spectra and other physical and chemical properties of inosine were markedly similar to those of "sporogen," a previously described endogenous sporogenesis factor present in sporulating Bacillus species.