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Satish T Pote,1,2 Mahesh S Sonawane,1 Praveen Rahi,1 Sunil R Shah,3 Yogesh S Shouche,1 Milind S Patole,1 Madhuri R Thakar,2 Rohit Sharma1 1National Centre for Microbial Resource (NCMR), National Centre for Cell Science, NCCS Complex, S.P. Pune University, Pune 411 007, Maharashtra, India; 2National AIDS Research Institute, Pune 411026, Maharashtra, India; 3Bharati Vidyapeeth Deemed University Medical College, Bharati Vidyapeeth, Pune 411043, Maharashtra, IndiaCorrespondence: Rohit SharmaNational Centre for Microbial Resource (NCMR), National Centre for Cell Science, NCCS Complex, S.P. Pune University, Ganeshkhind, Pune 411 007, Maharashtra, IndiaTel +91-20-25329035Fax +91-20-25692259Email rohit@nccs.res.inIntroduction: Species of genus Candida are part of the common microbiota of humans; however, some of the Candida species are known opportunistic pathogens. Formation of biofilms, resistance to antifungal drugs, and increase in asymptomatic infections demands more studies on isolation, identification and characterization of Candida from clinical samples.Methods: The present manuscript deals with assessment of authentic yeast identification by three methods viz., DNA sequencing of 28S rRNA gene, protein profiles using MALDI-TOF MS, and colony coloration on chromogenic media. Antifungal susceptibility and in vitro cell invasion assays were performed to further characterize these isolates.Results: Comparison of three methods showed that DNA sequence analysis correctly identified more than 99.4% of the isolates up to species level as compared to 89% by MALDI-TOF MS. In this study, we isolated a total of 176 yeasts from clinical samples and preliminary morphological characters indicated that these yeast isolates belong to the genus Candida. The species distribution of isolates was as follows: 75 isolates of Candida albicans (42.61%), 50 of C. tropicalis (28.40%), 22 of C. glabrata (12.5%), 14 of C. parapsilosis (7.95%) and 4 of Clavispora lusitaniae (2.27%). Other species like Cyberlindnera fabianii, Issatchenkia orientalis, Kluyveromyces marxianus, Kodamaea ohmeri, Lodderomyces sp., and Trichosporon asahii were less than 2%. Antifungal susceptibility assay performed with 157 isolates showed that most of the isolates were resistant to the four azoles viz., clotrimazole, fluconazole, itraconazole, and ketoconazole, and the frequency of resistance was more in non-albicans Candida isolates. The susceptibility to azole drugs ranged from 7% to 48%, while 75% of the tested yeasts were susceptible to nystatin. Moreover, 88 isolates were also tested for their capacity to invade human cells using HeLa cells. In vitro invasion assay showed that most of the C. albicans isolates showed epithelial cell invasion as compared to isolates belonging to C. glabrata, C. parapsilosis and C. tropicalis.Discussion: The identification of yeasts of clinical origin by sequencing of 28S rRNA gene performed better than MALDI-TOF MS. The present study reiterates the world scenario wherein there is a shift from Candida strains to emerging opportunistic pathogens which were earlier regarded as environmental strains. The present study enlightens the current understanding of identification methods for clinical yeast isolates, increased antifungal drug resistance, epithelial cell invasion as a virulence factor, and diversity of yeasts in Indian clinical samples.Keywords: yeast clinical isolates, chromogenic media, DNA sequencing, MALDI-TOF MS, antifungal drug resistance

The rapid improvements in next-generation sequencing technologies have made it possible to quickly access in-depth genome sequence data. This has resulted in a flurry of genome sequences of various organisms being published and made publicly available in the last two decades. However, not all organisms have genome sequence data available. Various factors play a role, such as the importance of the organism, either medically or economically, and the genome complexity of the organisms. Phlebotomus papatasi is the sandfly vector for the Leishmania parasite, which is the causative agent for leishmaniasis. P. papatasi is a hematophagous vector, and the female flies feed on human blood to complete their reproductive cycle. The P. papatasi genome is currently being sequenced as part of a multicentric consortium, and the genome sequence is not published to date. Hence, efforts to map its global proteome are hindered in P. papatasi. In such cases, comparative proteogenomic approaches can help map the global proteome of an unsequenced organism using homology-based methods., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Identifying new genes involved in virulence and drug resistance may hold the key to a better understanding of parasitic diseases. The proteogenomic profiling of various Leishmania species, the causative agents of leishmaniasis, has identified several novel genes, N- and C-terminal extensions of proteins, and corrections of existing gene models. Various virulence factors (VFs) responsible for leishmaniasis have been previously annotated through a proteogenomic approach, including the C-terminal extension of heat shock protein 70 (HSP70). Furthermore, the diversity of VFs across Leishmania donovani, L. infantum, L. major, and L. mexicana was determined using phylogenetic analysis. Moreover, protein-protein interaction networks (PPINs) of VFs with HSPs aid in making significant biological interpretations. Overall, an integrated omics approach involving proteogenomics was used to identify and study the relationship among VFs with other interacting proteins, including HSPs. This chapter provides a step-by-step guide to the identification of new genes in Leishmania using a proteogenomic approach and their functional assignment using a bioinformatics-based approach., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Although skin is the primary affected organ in Leprosy, the role of the skin microbiome in its pathogenesis is not well understood. Recent reports have shown that skin of leprosy patients (LP) harbours perturbed microbiota which grants inflammation and disease progression. Herein, we present the results of nested Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) which was initially performed for investigating the diversity of bacterial communities from lesional skin (LS) and non-lesional skin (NLS) sites of LP (n = 11). Further, we performed comprehensive analysis of 16S rRNA profiles corresponding to skin samples from participants (n = 90) located in two geographical locations i.e. Hyderabad and Miraj in India. The genus Staphylococcus was observed to be one of the representative bacteria characterizing healthy controls (HC; n = 30), which in contrast was underrepresented in skin microbiota of LP. Taxa affiliated to phyla Firmicutes and Proteobacteria were found to be signatures of HC and LS, respectively. Observed diversity level changes, shifts in core microbiota, and community network structure support the evident dysbiosis in normal skin microbiota due to leprosy. Insights obtained indicate the need for exploring skin microbiota modulation as a potential therapeutic option for leprosy.

Determining variations in protein abundance and/or posttranslational modification as a function of time or upon induction by a signal in a particular cell type is central to quantitative proteomics. Isobaric labeling methodologies now allow for parallel quantification of proteins at various conditions concurrently or multiplexing in relatively quantitative proteomics workflows. Hence, mapping the protein expression profiles of various developmental stages of Leishmania parasites is possible with high-resolution mass spectrometry. To analyze global changes in protein expression and cellular signaling pathways during Leishmania differentiation and development is possible with a quantitative proteomics approach. The tandem mass tags (TMT) approach provides a chemical labeling method based on the principle of amine reactive tags; the maximum number of conditions that can be multiplexed is 10-plex. We describe herein a detailed method for sample preparation, TMT-labeling, mass spectrometry and data analysis of different developmental stages of Leishmania donovani parasites. This quantitative proteomic approach is useful to study dynamic changes in protein expression levels during L. donovani differentiation, and also allows in-depth analysis of signaling pathways via phosphoproteomics.

Leprosy is an infectious disease that has predilection in skin and peripheral nerves. Skin has its own microbiome, however it is not extensively studied in Indian leprosy patients. Here, by using next-generation 16S rDNA sequencing, we have attempted to assess the skin associated microbial diversity pertaining to affected and unaffected skin of Indian leprosy patients. A total of 90 skin swab samples were collected from 60 individuals (30 healthy controls, 30 patients) residing in Hyderabad and Miraj, two distinct geographical locations in India to assess the homo/heterogeneity of skin microbial signatures. While a large increase in genus Methylobacterium and Pseudomonas was seen in patients from Miraj and Hyderabad respectively, a considerable decrease in genus Staphylococcus in the leprosy patients (as compared to controls) from both geographical locations was also observed. We expect that, these datasets can not-only provide further interesting insights, but will also help to observe dynamics of microbiome in the diseased state and generate hypotheses to test for skin microbiome transplantation studies in leprosy.

Idli, a naturally fermented Indian food, is prepared from a mixture of rice and black gram (lentil). To understand its microbial community during fermentation, detailed analysis of the structural and functional dynamics of the idli microbiome was performed by culture-dependent and -independent approaches. The bacterial diversity and microbial succession were assessed at different times of fermentation by 16S rRNA amplicon sequencing. Results highlighted that most microbiota belonged to phylum Firmicutes (70%) and Proteobacteria (22%). Denaturing gradient gel electrophoresis (DGGE) and quantitative PCR (qPCR) analysis confirmed the diversity and succession involved therein. A culture-dependent approach revealed that the microbially diverse populations were conserved across different geographical locations. The fermentation was primarily driven by lactic acid bacteria as they constitute 86% of the total bacterial population, and genus Weissella emerged as the most important organism in fermentation. The natural microbiota of the grains mainly drives the fermentation, as surface sterilized grains did not show any fermentation. Growth kinetics of idli microbiota and physicochemical parameters corroborated the changes in microbial dynamics, acid production, and leavening occurring during fermentation. Using a metagenomic prediction tool, we found that the major metabolic activities of these microbial fermenters were augmented during the important phase of fermentation. The involvement of the heterofermentative hexose monophosphate (HMP) pathway in batter leavening was substantiated by radiolabeled carbon dioxide generated from d-[1- 14 C]-glucose. Hydrolases degrading starch and phytins and the production of B vitamins were reported. Moreover, culturable isolates showing beneficial attributes, such as acid and bile tolerance, hydrophobicity, antibiotic sensitivity, and antimicrobial activity, suggest idli to be a potential dietary supplement. IMPORTANCE This is a comprehensive analysis of idli fermentation employing modern molecular tools which provided valuable information about the bacterial diversity enabling its fermentation. The study has demonstrated the relationship between the bacterial population and its functional role in the process. The nature of idli fermentation was found to be more complex than other food fermentations due to the succession of the bacterial population. Further studies using metatranscriptomics and metabolomics may enhance the understanding of this complex fermentation process. Moreover, the presence of microorganisms with beneficial properties plausibly makes idli a suitable functional food., (Copyright © 2019 American Society for Microbiology.)

Onychomycosis is the most common infection of the toe-nails or finger-nails and it may be caused by a large variety of fungal species. Achaetomium species which belong to the phylum Ascomycota (Family Chaetomiaceae), are usually soil saprophytes or endophytic fungi which have been rarely reported as human or animal pathogens. Here, we report a case of onychomycosis caused by Achaetomium strumarium in a healthy person who showed involvement of all fingers of both hands with yellowish brown discoloration. The causative agent isolated was identified as Achaetomium species by morphology, colony morphometry and growth at high temperature and as A. strumarium from DNA sequence of ITS region. Onychomycosis from this case responded satisfactorily with per os (P. O.; oral) and topical application of Terbinafine., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

[This corrects the article DOI: 10.18632/oncotarget.25717.].

Plant originated drugs/formulations are extensively prescribed by the physicians as a complementary therapy for treating various human ailments including cancer. In this study Prosopis juliflora leaves methanol extract was prepared and exposed to human breast cancer cell lines i.e. MDA-MB-231 and MCF-7 and human keratinocytes HaCaT as a representative of normal cells. Initially, a series of in vitro experiments like cell proliferation, migration, colony formation, cell cycle arrest and inhibition of angiogenesis. After confirmation of the efficient and selective activity against triple negative breast cancer cell line, we further evaluated the possible mechanism of inducing cell death and experiments like detection of reactive oxygen species, caspases and poly (ADP-ribose) polymerase cleavage study and Annexin V assay were performed. We also evaluated in vivo anti tumorigenic activity of the P. juliflora leaves by using 4T1 cells (a triple negative mouse origin breast cancer cell line) and BALB/c xenograft mouse model. In vitro experiments revealed that methanol extract of Prosopis juliflora leaves possess impressive anti-breast cancer activity more specifically against triple negative breast cancer cells, while the in vivo studies demonstrated that P. juliflora leaves extract significantly suppressed the 4T1 induced tumor growth. Present investigations clearly focus the significance of P. juliflora as an important resource for finding novel leads against triple negative breast cancer. The results may also act as a ready reference towards developing P. juliflora based formulation as an alternative and complementary medicine for the management of breast cancer., Competing Interests: CONFLICTS OF INTEREST The authors declare that there are no conflicts of interest to disclose.

Colletotrichum species have been reported infrequently as the cause of keratitis or subcutaneous lesions. The patient we describe developed keratitis after ocular trauma. The sample from the corneal scrapings grew Colletotrichum gloeosporioides as identified from morphological characters and DNA sequence of the 'Internal Transcribed Spacer' (ITS) region. The patient underwent topical application of amphotericin-B followed by itraconazole and natamycin treatment. Simultaneous oral voriconazole regimen leads to complete regression of corneal ulcer. This report highlights the fact that early and accurate identification and therapy can resolve keratitis caused by rare pathogen C. gloeosporioides., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)

Cutaneous leishmaniasis (CL) is caused by a kinetoplastid protozoan parasite Leishmania major, as a skin ulcer at the site of the sandfly bite. CL is curable and in most cases ulcers heal spontaneously within three to six months leaving a scar and disfiguration. Complete genome of L. major was reported in 2005 at the very initial phase of kinetoplastid parasite genome sequencing project. Presently, L. major genome is most studied and comprehensively annotated genome and therefore, it is being used as a reference genome for annotating recently sequenced Leishmanial genomes. A recent study reporting global transcriptome of L. major promastigotes, identified 1884 uniquely expressed non-coding RNAs (ncRNA) in L. major. In the current study, an in-depth analysis of the 1884 novel ncRNAs was carried out using a proteogenomic approach to identify their protein coding potential. Our analysis resulted in identification of eight novel protein coding genes based on mass spectrometry data. We have analyzed each of these eight novel CDS and in the process have improved the genome annotation of L. major on the basis of mass spectrometry derived peptide data. Although sequenced a decade ago, the improvement in the L. major genome annotation thus is an ongoing process., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Dengue viruses are transmitted to humans through the bites of infected female aedine mosquitoes. Differences in the composition and structure of bacterial communities in the midguts of mosquitoes may affect the vector's ability to transmit the disease. To investigate and analyse the role of midgut bacterial communities in viral transmission, midgut bacteria from three species, namely Stegomyia aegypti (= Aedes aegypti), Fredwardsius vittatus (= Aedes vittatus) and Stegomyia albopicta (= Aedes albopictus) (all: Diptera: Culicidae), from dengue-endemic and non-endemic areas of Rajasthan, India were compared. Construction and analyses of six 16S rRNA gene libraries indicated that Serratia spp.-related phylotypes dominated all clone libraries of the three mosquito species from areas in which dengue is not endemic. In dengue-endemic areas, phylotypes related to Aeromonas, Enhydrobacter spp. and uncultivated bacterium dominated the clone libraries of S. aegypti, F. vittatus and S. albopicta, respectively. Diversity indices analysis and real-time TaqMan polymerase chain reaction assays showed bacterial diversity and abundance in the midguts of S. aegypti to be higher than in the other two species. Significant differences observed among midgut bacterial communities of the three mosquito species from areas in which dengue is and is not endemic, respectively, may be related to the vectorial capacity of mosquitoes to carry dengue viruses and, hence, to the prevalence of disease in some areas., (© 2016 The Royal Entomological Society.)

Objective: The occurrence of yeast infections in humans has increased, with the species belonging to genus Candida still being the most common cause of infection. Nevertheless, infections caused by less common yeasts have been widely reported in recent years. The main objective of this study was to assess the potential of these less common saprophytic yeasts to invade the host cell, which is essential for causing systemic infections., Material and Methods: Various yeast isolates were identified by DNA sequence information of PCR amplified ITS region. The purported saprophytic yeasts were characterized for internalization by mammalian cells in vitro, by staining the F-actin., Conclusion: The identification of different yeast isolates from various patients revealed that 70% of the isolates belonged to the genus Candida, while remaining 30% of the isolates were yeasts not belonging to genus Candida. These non-Candida clinical isolates, either in yeast or hyphal forms, were efficiently internalized by human epithelial cells. The internalization was marked by a process of actin polymerization surrounding the invading yeast. Such uptake by epithelial cells signifies traversal of cell barrier by yeast cells during infection in vivo., (Copyright © 2015 Elsevier Masson SAS. All rights reserved.)

The debilitating disease kala-azar or visceral leishmaniasis is caused by the kinetoplastid protozoan parasite Leishmania donovani. The parasite is transmitted by the hematophagous sand fly vector of the genus Phlebotomus in the old world and Lutzomyia in the new world. The predominant Phlebotomine species associated with the transmission of kala-azar are Phlebotomus papatasi and Phlebotomus argentipes. Understanding the molecular interaction of the sand fly and Leishmania, during the development of parasite within the sand fly gut is crucial to the understanding of the parasite life cycle. The complete genome sequences of sand flies (Phlebotomus and Lutzomyia) are currently not available and this hinders identification of proteins in the sand fly vector. The current study utilizes a three frame translated transcriptomic data of P. papatasi in the absence of genomic sequences to analyze the mass spectrometry data of P. papatasi cell line using a proteogenomic approach. Additionally, we have carried out the proteogenomic analysis of P. papatasi by comparative homology-based searches using related sequenced dipteran protein data. This study resulted in the identification of 1313 proteins from P. papatasi based on homology. Our study demonstrates the power of proteogenomic approaches in mapping the proteomes of unsequenced organisms., (Copyright © 2015 Elsevier B.V. All rights reserved.)

The recent 2013-15 epidemic of Ebola virus disease (EVD) has initiated extensive sequencing and analysis of ebolavirus genomes. All ebolavirus genomes available until December 2014 have been collated and analyzed in this study to obtain phylogenetic relationship and uncover the variations amongst them. The terminal 'leader' and 'trailer' nucleotide sequences of the genomes were omitted and analysis of the intermediate region accommodating the sole seven genes (hepta-CDS region) of the virus showed relative stability of the genome, including the ones isolated from the current epidemic. The genome information was scrutinized to detect the variation in the surface glycoprotein gene and annotate its three protein products, resulting from its atypical transcription. This study will make an easy understanding of the genomes for those who desire to exploit the genome sequences for different investigations in EVD., (Copyright © 2015 Elsevier Inc. All rights reserved.)

There is a growing interest in subtype (ST) analysis of the intestinal parasite Blastocystis due to its extensive genetic diversity that might reflect differences in pathogenicity. Although essential for reference, few studies are available on Blastocystis in healthy individuals. Moreover, molecular epidemiology data on Blastocystis in India still remain to emerge. In the present study we identified the prevalence and ST distribution of Blastocystis in healthy Indian individuals. A total of 220 stool samples were obtained; four of 100 samples from 100 adults were chosen randomly for construction of small subunit (SSU) rRNA gene clone libraries in order to elucidate micro-eukaryotic diversity in the human gut. From the SSU rDNA library, 64 sequences annotated to Blastocystis were used for ST analysis along with sequences obtained by direct sequencing of SSU rDNA PCR products amplified from the remaining samples and generated using primers targeting Blastocystis. Of 220 stool samples collected, 120 samples from 30 infants (aged 1week to 1year) were PCR-negative. Of the remaining 100 samples from 100 adults, 27 resulted in specific amplification. Out of these 27, four samples were suspected of mixed ST infection and so these samples were further analyzed by construction of clone libraries. Analysis of cloned sequences revealed that indeed 2 samples had mixed ST infection (ST1 and ST3) while the remaining two showed infection with two separate ST3 strains. ST3 was the most common ST present in our study group (100%) followed by ST1 (7.4%); ST1 was seen only in mixed infections. SSU rDNA clone library sequences generated by processing of pooled samples were identified as ST3. The majority of ST3 sequences exhibited allele 34 commonly found in the European population., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Leishmania donovani is a kinetoplastid protozoan that causes a severe and fatal disease kala-azar, or visceral leishmaniasis. L. donovani infects human host after the phlebotomine sandfly takes a blood meal and resides within the phagolysosome of infected macrophages. Previous studies on host-parasite interactions have not focused on Leishmania organelles and the role that they play in the survival of this parasite within macrophages. Leishmania possess glycosomes that are unique and specialized subcellular microbody organelles. Glycosomes are known to harbor most peroxisomal enzymes and, in addition, they also possess nine glycolytic enzymes. In the present study, we have carried out proteomic profiling using high resolution mass spectrometry of a sucrose density gradient-enriched glycosomal fraction isolated from L. donovani promastigotes. This study resulted in the identification of 4022 unique peptides, leading to the identification of 1355 unique proteins from a preparation enriched in L. donovani glycosomes. Based on protein annotation, 566 (41.8%) were identified as hypothetical proteins with no known function. A majority of the identified proteins are involved in metabolic processes such as carbohydrate, lipid, and nucleic acid metabolism. Our present proteomic analysis is the most comprehensive study to date to map the proteome of L. donovani glycosomes.

In past years, nanostructured lipid carriers (NLCs) have emerged as novel topical antioxidant delivery systems because of combined positive features of liposomes and polymeric nanoparticles. Here, we seek to unlock the possibility of idebenone (IDB; an antioxidant)-loaded NLCs (IDB-NLCs) cellular interactions such as, viability and uptake, and its photoprotective effects against Ultraviolet-B (UVB)-mediated oxidative stress in immortal human keratinocyte cell line (HaCaT). The two-step preformulation strategy followed by three-level, three-variable, L9 (3(3)) Taguchi robust orthogonal design employed was important in improving IDB-NLCs key physicochemical aspects such as, entrapment efficiency, drug release (sustained), occlusion, skin deposition and physical stability. UV crosslinker, confocal microscopy and flow cytometry techniques were used to (1) mediate oxidative stress in HaCaT cells, (2) study a qualitative cellular uptake, (3) measure intracellular reactive oxygen species (ROS), and mitochondrial membrane potential, respectively. NLCs markedly improved biocompatibility of IDB under normal as well as stress conditions. Quantitative and qualitative cell uptake studies demonstrated a significant uptake of IDB-NLCs (3-fold increase) and nile red-labeled IDB-NLCs (NR-IDB-NLCs) at 2 h, respectively, hence exerted improved photoprotective effects., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Leishmania donovani is a kinetoplastid protozoan parasite which causes the fatal disease visceral leishmaniasis in humans. Genome sequencing of L. donovani revealed information about the arrangement of genes and genome architecture. After curation of the genome sequence, many genes in L. donovani were assigned as truncated or "partial" genes by the genome sequencing group. In the present study, we have carried out an extensive analysis and attempted to improve the gene models of these partial genes. Our analysis resulted in the identification of 308 partial genes in L. donovani, which were further categorized as C-terminal extensions, joining of genes, tandemly repeated paralogs and wrong chromosomal assignments. We have analyzed each of these genes from these categories and have improved the annotation of existing gene models in L. donovani. Some of these corrections have been confirmed by mass spectrometry derived peptide data from our previous comparative proteogenomics study in L. donovani., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: Oral therapy for pulmonary tuberculosis (TB) treatment suffers from the limitation of hepatic metabolism leading insufficient concentration of antitubercular (anti-TB) drugs in alveolar macrophage which harbors Mycobacterium tuberculosis (MTB). Targeted aerosol delivery of antituberculous drug to lung is efficient for treating local lung TB infection., Objective: The present study was aimed to evaluate rifapentine (RPT) loaded proliposomal dry powder for inhalation (RLDPI) for anti-TBactivity and cytotoxicity in vitro. In vivo toxicity study was also undertaken in Wistar rats to determine safe concentration of RLDPI for administration., Materials and Methods: Anti-TB activity of developed RLDPI was assessed using drug susceptibility testing (DST) on Mycobacteria growth indicator tube (MGIT) method. In vitro cytotoxicity was performed in A549 cell lines and IC50 values were used to compare the cytotoxicity of formulation with pure RPT. In vivo repeated dose toxicity study was undertaken using Wistar rats at three different doses for 28-days by intratracheal insufflations method., Results: The results of DST study revealed sensitivity of tubercle bacteria to RLDPI at concentration equivalent to 10 μg/mL of RPT. This study confirmed anti-TB potential of RPT in spray-dried RLDPI, though the spray drying method is reported to reduce activity of drugs. Cytotoxicity study in A549 cells demonstrated that RPT when encapsulated in liposomes as RLDPI was safe to cells as compared to pure RPT. In vivo toxicity study revealed that RPT in the form of RLDPI was safe at 1 and 5 mg/kg dose. However, mortality was seen at higher dose (10 mg/kg), possibly because of liver and kidney damage., Conclusion: Thus, these studies demonstrated safety of RLDPI for the treatment of pulmonary TB.

Among the neglected tropical diseases, leishmaniasis is one of the most devastating, resulting in significant mortality and contributing to nearly 2 million disability-adjusted life years. Cutaneous leishmaniasis is a debilitating disorder caused by the kinetoplastid protozoan parasite Leishmania major, which results in disfiguration and scars. L. major genome was the first to be sequenced within the genus Leishmania. Use of proteomic data for annotating genomes is a complementary approach to conventional genome annotation approaches and is referred to as proteogenomics. We have used a proteogenomics-based approach to map the proteome of L. major and also annotate its genome. In this study, we searched L. major promastigote proteomic data against the annotated L. major protein database. Additionally, we searched the proteomic data against six-frame translated L. major genome. In all, we identified 3613 proteins in L. major promastigotes, which covered 43% of its proteome. We also identified 26 genome search-specific peptides, which led to the identification of three novel genes previously not identified in L. major. We also corrected the annotation of N-termini of 15 genes, which resulted in extension of their protein products. We have validated our proteogenomics findings by RT-PCR and sequencing. In addition, our study resulted in identification of 266 N-terminally acetylated peptides in L. major, one of the largest acetylated peptide datasets thus far in Leishmania. This dataset should be a valuable resource to researchers focusing on neglected tropical diseases.

The kinetoplastid protozoan parasite, Leishmania donovani, is the causative agent of kala azar or visceral leishmaniasis. Kala azar is a severe form of leishmaniasis that is fatal in the majority of untreated cases. Studies on proteomic analysis of L. donovani thus far have been carried out using homology-based identification based on related Leishmania species (L. infantum, L. major and L. braziliensis) whose genomes have been sequenced. Recently, the genome of L. donovani was fully sequenced and the data became publicly available. We took advantage of the availability of its genomic sequence to carry out a more accurate proteogenomic analysis of L. donovani proteome using our previously generated dataset. This resulted in identification of 17,504 unique peptides upon database-dependent search against the annotated proteins in L. donovani. These peptides were assigned to 3999 unique proteins in L. donovani. 2296 proteins were identified in both the life stages of L. donovani, while 613 and 1090 proteins were identified only from amastigote and promastigote stages, respectively. The proteomic data was also searched against six-frame translated L. donovani genome, which led to 255 genome search-specific peptides (GSSPs) resulting in identification of 20 novel genes and correction of 40 existing gene models in L. donovani., Biological Significance: Leishmania donovani genome sequencing was recently completed, which permitted us to use a proteogenomic approach to map its proteome and to carry out annotation of it genome. This resulted in mapping of 50% (3999 proteins) of L. donovani proteome. Our study identified 20 novel genes previously not predicted from the L. donovani genome in addition to correcting annotations of 40 existing gene models. The identified proteins may help in better understanding of stage-specific protein expression profiles in L. donovani and to identify novel stage-specific drug targets in L. donovani which could be used in the treatment of leishmaniasis. This article is part of a Special Issue entitled: Trends in Microbial Proteomics., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Background: Modulated immune signal (CD14-TLR and TNF) in leishmaniasis can be linked to EGFR pathway involved in wound healing, through crosstalk points. This signaling network can be further linked to a synthetic gene circuit acting as a positive feedback loop to elicit a synchronized intercellular communication among the immune cells which may contribute to a better understanding of signaling dynamics in leishmaniasis., Methods: Network reconstruction with positive feedback loop, simulation (ODE 15s solver) and sensitivity analysis of CD14-TLR, TNF and EGFR was done in SimBiology (MATLAB 7.11.1). Cytoscape and adjacency matrix were used to calculate network topology. PCA was extracted by using sensitivity coefficient in MATLAB. Model reduction was done using time, flux and sensitivity score., Results: Network has five crosstalk points: NIK, IκB-NFκB and MKK (4/7, 3/6, 1/2) which show high flux and sensitivity. PI3K in EGFR pathway shows high flux and sensitivity. PCA score was high for cytoplasmic ERK1/2, PI3K, Atk, STAT1/3 and nuclear JNK. Of the 125 parameters, 20% are crucial as deduced by model reduction., Conclusions: EGFR can be linked to CD14-TLR and TNF through the MAPK crosstalk points. These pathways may be controlled through Ras and Raf that lie upstream of signaling components ERK ½ (c) and JNK (n) that have a high PCA score via a synthetic gene circuit for activating cell-cell communication to elicit an inflammatory response. Also a disease resolving effect may be achieved through PI3K in the EGFR pathway., General Significance: The reconstructed signaling network can be linked to a gene circuit with a positive feedback loop, for cell-cell communication resulting in synchronized response in the immune cell population, for disease resolving effect in leishmaniasis., (© 2013 Elsevier B.V. All rights reserved.)

Network of signaling proteins and functional interaction between the infected cell and the leishmanial parasite, though are not well understood, may be deciphered computationally by reconstructing the immune signaling network. As we all know signaling pathways are well-known abstractions that explain the mechanisms whereby cells respond to signals, collections of pathways form networks, and interactions between pathways in a network, known as cross-talk, enables further complex signaling behaviours. In silico perturbations can help identify sensitive crosstalk points in the network which can be pharmacologically tested. In this study, we have developed a model for immune signaling cascade in leishmaniasis and based upon the interaction analysis obtained through simulation, we have developed a model network, between four signaling pathways i.e., CD14, epidermal growth factor (EGF), tumor necrotic factor (TNF) and PI3 K mediated signaling. Principal component analysis of the signaling network showed that EGF and TNF pathways can be potent pharmacological targets to curb leishmaniasis. The approach is illustrated with a proposed workable model of epidermal growth factor receptor (EGFR) that modulates the immune response. EGFR signaling represents a critical junction between inflammation related signal and potent cell regulation machinery that modulates the expression of cytokines.

Differences in midgut bacterial communities of Aedes aegypti, the primary mosquito vector of dengue viruses (DENV), might influence the susceptibility of these mosquitoes to infection by DENV. As a first step toward addressing this hypothesis, comparative analysis of bacterial communities from midguts of mosquito strains with differential genetic susceptibility to DENV was performed. 16S rRNA gene libraries and real-time PCR approaches were used to characterize midgut bacterial community composition and abundance in three Aedes aegypti strains: MOYO, MOYO-R, and MOYO-S. Although Pseudomonas spp.-related clones were predominant across all libraries, some interesting and potentially significant differences were found in midgut bacterial communities among the three strains. Pedobacter sp.- and Janthinobacterium sp.-related phylotypes were identified only in the MOYO-R strain libraries, while Bacillus sp. was detected only in the MOYO-S strain. Rahnella sp. was found in MOYO-R and MOYO strains libraries but was absent in MOYO-S libraries. Both 16S rRNA gene library and real-time PCR approaches confirmed the presence of Pedobacter sp. only in the MOYO-R strain. Further, real-time PCR-based quantification of 16S rRNA gene copies showed bacterial abundance in midguts of the MOYO-R strain mosquitoes to be at least 10-100-folds higher than in the MOYO-S and MOYO strain mosquitoes. Our study identified some putative bacteria with characteristic physiological properties that could affect the infectivity of dengue virus. This analysis represents the first report of comparisons of midgut bacterial communities with respect to refractoriness and susceptibility of Aedes aegypti mosquitoes to DENV and will guide future efforts to address the potential interactive role of midgut bacteria of Aedes aegypti mosquitoes in determining vectorial capacity for DENV.

Germ band retraction (GBR) stage is one of the important stages during insect development. It is associated with an extensive epithelial morphogenesis and may also be pivotal in generation of morphological diversity in insects. Despite its importance, only a handful of studies report the transcriptome repertoire of this stage in insects. Here, we report generation, annotation and analysis of ESTs from the embryonic stage (16-22 h post fertilization) of laboratoryreared Anopheles stephensi mosquitoes. A total of 1002 contigs were obtained upon clustering of 1140 high-quality ESTs, which demonstrates an astonishingly low transcript redundancy (12.1 percent). Putative functions were assigned only to 213 contigs (21 percent), comprising mainly of transcripts encoding protein synthesis machinery. Approximately 78 percent of the transcripts remain uncharacterized, illustrating a lack of sequence information about the genes expressed in the embryonic stages of mosquitoes. This study highlights several novel transcripts, which apart from insect development, may significantly contribute to the essential biological complexity underlying insect viability in adverse environments. Nonetheless, the generated sequence information from this work provides a comprehensive resource for genome annotation, microarray development, phylogenetic analysis and other molecular biology applications in entomology.

Kutajarista is an Ayurvedic fermented herbal formulation prescribed for gastrointestinal disorders. This herbal formulation undergoes a gradual fermentative process and takes around 2 months for production. In this study, microbial composition at initial stages of fermentation of Kutajarista was assessed by culture independent 16S rRNA gene clone library approach. Physicochemical changes were also compared at these stages of fermentation. High performance liquid chromatography-mass spectrometry analysis showed that Gallic acid, Ellagic acid, and its derivatives were the major chemical constituents recovered in this process. At 0 day of fermentation, Lactobacillus sp., Acinetobacter sp., Alcaligenes sp., and Methylobacterium sp. were recovered, but were not detected at 8 day of fermentation. Initially, microbial diversity increased after 8 days of fermentation with 11 operational taxonomic units (OTUs), which further decreased to 3 OTUs at 30 day of fermentation. Aeromonas sp., Pseudomonas sp., and Klebsiella sp. dominated till 30 day of fermentation. Predominance of γ- Proteobacteria and presence of gallolyl derivatives at the saturation stage of fermentation implies tannin degrading potential of these microbes. This is the first study to highlight the microbial role in an Ayurvedic herbal product fermentation.

Bacillus lentus BI377 (B. lentus BI377) an alkaliphilic strain has accomplished the discriminate color removal strategy for Reactive Red sulfonated azoic recalcitrant irrespective of their molecular structure. During the decolorization experiment, it was observed that the diazo dye first followed chromophoric cleavage by azoreductase via typical azoreduction whereas, in case of monoazo dye, cleavage took place by peroxidase via successive electron transfers to oxide surface resulting in the asymmetric cleavage of the azo bond. Dismutation of oxidative stress by reactive metabolites has confirmed by superoxide dismutase activity. Carbon monoxide (CO) binding spectra, the content of cytochrome P450 and spectroscopy analysis by GCMS, FTIR and (1)H NMR of intermediate metabolites indicated the differentiate pattern of diazo and monoazo dye decolorization fuse to central metabolic pathway. Declined percentage of TOC and the cytotoxicity (MTT) study confirmed that environmentally benign intermediates may lead to mineralization., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

In this study fecal microflora of human infants born through vaginal delivery (VB) and through cesarean section (CB) were investigated using culture-independent 16S rDNA cloning and sequencing approach. The results obtained clearly revealed that fecal microbiota of VB infants distinctly differ from those in their counterpart CB infants. The intestinal microbiota of infants delivered by cesarean section appears to be more diverse, in terms of bacteria species, than the microbiota of vaginally delivered infants. The most abundant bacterial species present in VB infants were Acinetobacter sp., Bifidobacterium sp. and Staphylococcus sp. However, CB infant's fecal microbiota was dominated with Citrobacter sp., Escherichia coli and Clostridium difficile. The intestinal microbiota of cesarean section delivered infants in this study was also characterized by an absence of Bifidobacteria species. An interesting finding of our study was recovery of large number of Acinetobacter sp. consisting of Acinetobacter pittii (former Acinetobacter genomic species 3), Acinetobacter junii and Acinetobacter baumannii in the VB infants clone library. Among these, Acinetobacter baumannii is a known nosocomial pathogen and Acinetobacter pittii (genomic species 3) is recently recognized as clinically important taxa within the Acinetobacter calcoaceticus-Acinetobacter baumannii (ACB) complex. Although none of the infants had shown any sign of clinical symptoms of disease, this observation warrants a closer look.

The gastrointestinal (GI) tract of invasive land snail Achatina fulica is known to harbor metabolically active bacterial communities. In this study, we assessed the bacterial diversity in the different regions of GI tract of Giant African snail, A. fulica by culture-independent and culture-dependent methods. Five 16S rRNA gene libraries from different regions of GI tract of active snails indicated that sequences affiliated to phylum γ-Proteobacteria dominated the esophagus, crop, intestine, and rectum libraries, whereas sequences affiliated to Tenericutes dominated the stomach library. On phylogenetic analysis, 30, 27, 9, 27, and 25 operational taxonomic units (OTUs) from esophagus, crop, stomach, intestine, and rectum libraries were identified, respectively. Estimations of the total bacterial diversity covered along with environmental cluster analysis showed highest bacterial diversity in the esophagus and lowest in the stomach. Thirty-three distinct bacterial isolates were obtained, which belonged to 12 genera of two major bacterial phyla namely γ-Proteobacteria and Firmicutes. Among these, Lactococcus lactis and Kurthia gibsonii were the dominant bacteria present in all GI tract regions. Quantitative real-time polymerase chain reaction (qPCR) analysis indicated significant differences in bacterial load in different GI tract regions of active and estivating snails. The difference in the bacterial load between the intestines of active and estivating snail was maximum. Principal component analysis (PCA) of terminal restriction fragment length polymorphism suggested that bacterial community structure changes only in intestine when snail enters estivation state., (© 2012 The Authors. Published by Blackwell Publishing Ltd.)

We have carried out a polyphasic taxonomic characterization of Bacillus beijingensis DSM 19037(T) and Bacillus ginsengi DSM 19038(T), which are closely related phylogenetically to Bhargavaea cecembensis LMG 24411(T). All three strains are Gram-stain-positive, non-motile, moderately halotolerant and non-spore-forming. 16S rRNA gene sequence analyses showed that the strains constituted a coherent cluster, with sequence similarities between 99.7 and 98.7 %. The percentage similarity on the basis of amino acid sequences deduced from partial gyrB gene nucleotide sequences of these three type strains was 96.1-92.7 %. Phylogenetic trees based on the 16S rRNA gene and GyrB amino acid sequences, obtained by using three different algorithms, were consistent and showed that these three species constituted a deeply rooted cluster separated from the clades represented by the genera Bacillus, Planococcus, Planomicrobium, Sporosarcina, Lysinibacillus, Viridibacillus, Kurthia and Geobacillus, supporting their placement in the genus Bhargavaea. All three type strains have menaquinone MK-8 as the major respiratory quinone and showed similar fatty acid profiles. The main polar lipids present in the three type strains were diphosphatidylglycerol and phosphatidylglycerol, and the three strains showed peptidoglycan type A4α with L-lysine as the diagnostic diamino acid. The DNA G+C contents of Bacillus beijingensis DSM 19037(T), Bacillus ginsengi DSM 19038(T) and Bhargavaea cecembensis LMG 24411(T) were 53.1, 50.2 and 53.7 mol%, respectively. The level of DNA-DNA hybridization among the three strains was 57-39 %, indicating that they are members of different species of the genus Bhargavaea. The phenotypic data are consistent with the placement of these three species in a single genus and support their differentiation at the species level. On the basis of these data, we have emended the description of the genus Bhargavaea and propose the reclassification of Bacillus beijingensis and Bacillus ginsengi to the genus Bhargavaea, as Bhargavaea beijingensis comb. nov. (type strain ge10(T)  = DSM 19037(T)  = CGMCC 1.6762(T)) and Bhargavaea ginsengi comb. nov. (type strain ge14(T)  = DSM 19038(T)  = CGMCC 1.6763(T)).

Obesity is a consequence of a complex interplay between the host genome and the prevalent obesogenic factors among the modern communities. The role of gut microbiota in the pathogenesis of the disorder was recently discovered; however, 16S-rRNA-based surveys revealed compelling but community-specific data. Considering this, despite unique diets, dietary habits and an uprising trend in obesity, the Indian counterparts are poorly studied. Here, we report a comparative analysis and quantification of dominant gut microbiota of lean, normal, obese and surgically treated obese individuals of Indian origin. Representative gut microbial diversity was assessed by sequencing fecal 16S rRNA libraries for each group (n=5) with a total of over 3000 sequences. We detected no evident trend in the distribution of the predominant bacterial phyla, Bacteroidetes and Firmicutes. At the genus level, the bacteria of genus Bacteroides were prominent among the obese individuals, which was further confirmed by qPCR (P less than 0.05). In addition, a remarkably high archaeal density with elevated fecal SCFA levels was also noted in the obese group. On the contrary, the treated-obese individuals exhibited comparatively reduced Bacteroides and archaeal counts along with reduced fecal SCFAs. In conclusion, the study successfully identified a representative microbial diversity in the Indian subjects and demonstrated the prominence of certain bacterial groups in obese individuals; nevertheless, further studies are essential to understand their role in obesity.

The micro-eukaryotic diversity from the human gut was investigated using universal primers directed towards 18S rRNA gene, fecal samples being the source of DNA. The subjects in this study included two breast-fed and two formula-milk-fed infants and their mothers. The study revealed that the infants did not seem to harbour any microeukaryotes in their gut. In contrast, there were distinct eukaryotic microbiota present in the mothers. The investigation is the first of its kind in the comparative study of the human feces to reveal the presence of micro-eukaryotic diversity variance in infants and adults from the Indian subcontinent. The micro-eukaryotes encountered during the investigation include known gut colonizers like Blastocystis and some fungi species. Some of these micro-eukaryotes have been speculated to be involved in clinical manifestations of various diseases. The study is an attempt to highlight the importance of micro-eukaryotes in the human gut.

Members of the genus Wolbachia are intracellular bacteria that are widespread in arthropods and establish diverse symbiotic associations with their hosts, ranging from mutualism to parasitism. Here we present the first detailed analyses of Wolbachia in butterflies from India with screening of 56 species. Twenty-nine species (52%) representing five families were positive for Wolbachia. This is the first report of Wolbachia infection in 27 of the 29 species; the other two were reported previously. This study also provides the first evidence of infection in the family Papilionidae. A striking diversity was observed among Wolbachia strains in butterfly hosts based on five multilocus sequence typing (MLST) genes, with 15 different sequence types (STs). Thirteen STs are new to the MLST database, whereas ST41 and ST125 were reported earlier. Some of the same host species from this study carried distinctly different Wolbachia strains, whereas the same or different butterfly hosts also harbored closely related Wolbachia strains. Butterfly-associated STs in the Indian sample originated by recombination and point mutation, further supporting the role of both processes in generating Wolbachia diversity. Recombination was detected only among the STs in this study and not in those from the MLST database. Most of the strains were remarkably similar in their wsp genotype, despite divergence in MLST. Only two wsp alleles were found among 25 individuals with complete hypervariable region (HVR) peptide profiles. Although both wsp and MLST show variability, MLST gives better separation between the strains. Completely different STs were characterized for the individuals sharing the same wsp alleles.

We have checked the utility of DNA barcoding for species identification of nymphalid butterflies from Western Ghats of India by using 650 bp sequence of mitochondrial gene cytochrome c oxidase subunit I. Distinct DNA barcoding gap (i.e. difference between intraspecies and interspecies nucleotide divergence), exists between species studied here. When our sequences were compared with the sequences of the conspecifics submitted from different geographic regions, nine cases of deep intraspecies nucleotide divergences were observed. In spite of this, NJ (Neighbour Joining) clustering analysis successfully discriminated all species. Observed cases of deep intraspecies nucleotide divergences certainly warrant further study.

Visceral leishmaniasis or kala azar is the most severe form of leishmaniasis and is caused by the protozoan parasite Leishmania donovani. There is no published report on L. donovani genome sequence available till date, although the genome sequences of three related Leishmania species are already available. Thus, we took a proteogenomic approach to identify proteins from two different life stages of L. donovani. From our analysis of the promastigote (insect) and amastigote (human) stages of L. donovani, we identified a total of 22,322 unique peptides from a homology-based search against proteins from three Leishmania species. These peptides were assigned to 3711 proteins in L. infantum, 3287 proteins in L. major, and 2433 proteins in L. braziliensis. Of the 3711 L. donovani proteins that were identified, the expression of 1387 proteins was detectable in both life stages of the parasite, while 901 and 1423 proteins were identified only in promastigotes and amastigotes life stages, respectively. In addition, we also identified 13 N-terminally and one C-terminally extended proteins based on the proteomic data search against the six-frame translated genome of the three related Leishmania species. Here, we report results from proteomic profiling of L. donovani, an organism with an unsequenced genome., (© 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

House flies (Musca domestica L.) are cosmopolitan, ubiquitous, synanthropic insects that serve as mechanical or biological vectors for various microorganisms. To fully assess the role of house flies in the epidemiology of human diseases, it is essential to understand the diversity of microbiota harbored by natural fly populations. This study aimed to identify the diversity of house fly gut bacteria by both culture-dependent and culture-independent approaches. A total of 102 bacterial strains were isolated from the gut of 65 house flies collected from various public places including a garden, public park, garbage/dump area, public toilet, hospital, restaurant/canteen, mutton shop/market, and house/human habitation. Molecular phylogenetic analyses placed these isolates into 22 different genera. The majority of bacteria identified were known potential pathogens of the genera Klebsiella, Aeromonas, Shigella, Morganella, Providencia, and Staphylococcus. Culture-independent methods involved the construction of a 16S rRNA gene clone library, and sequence analyses supported culture recovery results. However, additional bacterial taxa not determined via culture recovery were revealed using this methodology and included members of the classes Alphaproteobacteria, Deltaproteobacteria, and the phylum Bacteroidetes. Here, we show that the house fly gut is an environmental reservoir for a vast number of bacterial species, which may have impacts on vector potential and pathogen transmission., (© 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.)

Four novel cell lines from tissues of eye, gill, kidney and brain of Etroplus suratensis were developed and characterized. The cell lines of eye, gill, kidney and brain were sub-cultured for 245, 185, 170 and 90 passages, respectively, since 2008. These cell lines showed predominantly epithelial-like cells. Effects of temperature and foetal bovine serum concentration on the growth of these cell lines were examined and optimum growth was found at the temperature of 28° C with 20% foetal bovine serum. All the four cell lines were successfully cryopreserved and revived at different passage levels. Cell-cycle analysis of these cell lines was carried out by fluorescence-activated cell sorting. Polymerase chain reaction (PCR) products obtained from the cells and tissues of E. suratensis with primers specific to the conserved region of 16S ribosomal RNA and cytochrome oxidase I genes of E. suratensis revealed the origin of cell lines from E. suratensis. Antibodies raised against the tissues and cells of eye, kidney and gill were highly cross reacted to their specific tissue and cells of E. suratensis. Chromosomal analysis revealed that E. suratensis cells have a normal diploid karyotype with 2n = 48. The cells of these cell lines were successfully transfected with pEGFP vector DNA. The eye (IEE), gill (IEG) and kidney (IEK) cell lines were found to be susceptible to nodavirus but resistant to infectious pancreatic necrosis virus (IPNV). The cells of gill, kidney and eye were applied to test the cytotoxicity of tannery effluents., (© 2012 The Authors. Journal of Fish Biology © 2012 The Fisheries Society of the British Isles.)

The present study deals with development of aerobic granules from textile wastewater sludge and challenged with different concentration of reactive blue 59 (RB59) to test their dye degradation potential. The granules efficiently degraded reactive blue 59 and also sustained higher dye loading of up to 5.0 g l(-1). The significant induction of enzymes azoreductase and cytochrome P-450 indicated their prominent role in the dye degradation while genotoxicity studies demonstrated that the biotransformed product of the dye as non-toxic. The microbial community of the textile dyes degrading aerobic sludge granules analyzed by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), revealed significantly diverse dye degrading microbial community belonging to alpha-, beta-, and gamma-proteobacteria., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Exposure of skin to various chemical and physical agents results in excessive stress to the outermost cell layer of the skin, causing different degenerative effects that can be minimized by using antioxidant formulations. The major challenge, in this regard, is to develop a formulation, which can prevent photodegradation of the actives, thus allowing a significant amount to be deposited at the site. In recent decades, liposomal formulations have been extensively employed to overcome the barrier properties of the skin and photodegradation of actives. In the present study, chitosan-reduced gold nanoparticles were investigated for its potential as a carrier to prepare liposomes by a spray-drying method. Liposomes so obtained were characterized for phospholipid recovery, diffuse reflectance infrared Fourier transform (DRIFT) spectroscopy, particle size, zeta potential, encapsulation efficiency, and deposition of drug and gold nanoparticles in the rat skin. Further, a liposomal gel formulation was prepared using Carbopol® 980 NF (Noveon Systems, Kochi, India) and evaluated for drug deposition in the skin. Antioxidant activity of vitamin C encapsulated in gold liposomes was determined on a human leukemia (HL-60) cell line. The use of gold nanoparticles as a carrier showed improved phospholipid recovery and thus overcomes the liposome scalability problem. DRIFT spectra confirmed the presence of phospholipid in the formulation. Liposomal gel showed improved drug deposition, as compared to control and marketed preparations. A more interesting contribution of the chitosan-reduced gold nanoparticles was an enhanced antioxidant activity seen in case of the vitamin C-loaded gold liposomal formulation. Liposomal formulation was found to be stable for 3 months at 30°C and 65% relative humidity.

A Gram-negative, facultatively anaerobic, rod-shaped, catalase- and oxidase-positive bacterium, motile by means of a single polar flagellum and designated strain KJW27(T), was isolated from the marine sediment of Karwar jetty, west coast of India. The strain was β-haemolytic and grew with 0-10 % (w/v) NaCl, at 10-45 °C and at pH 6.5-10, with optimum growth with 2 % (w/v) NaCl, at 37 °C and at pH 7.5. The major fatty acids were iso-C₁₅:₀ (22.2 %), C₁₇:₁ω8c (21 %), summed feature 3 (comprising C₁₆:₁ω7c and/or C₁₆:₁ω6c; 10.2 %), C₁₆:₀ (7.1 %), iso-C₁₃:₀ (5.6 %) and C₁₇:₀ (4.4 %). The DNA G+C content was 51.2 mol%. Phylogenetic analysis based on 16S rRNA and gyrB gene sequences showed that strain KJW27(T) forms a lineage within the genus Shewanella and is closely related to Shewanella algae ATCC 51192(T) (98.8 %), Shewanella haliotis DW01(T) (98.8 %) and Shewanella chilikensis JC5(T) (98.2 %). Sequence identity with other members of this genus ranges from 92.2 to 96.4 %. The DNA-DNA relatedness of strain KJW27(T) with S. algae ATCC 51192(T), S. haliotis DW01(T) and S. chilikensis JC5(T) was 52, 44 and 33 %, respectively. The phenotypic, genotypic and DNA-DNA relatedness data indicate that strain KJW27(T) should be distinguished from S. algae ATCC 51192(T), S. haliotis DW01(T) and S. chilikensis JC5(T). On the basis of the data presented in this study, strain KJW27(T) represents a novel species, for which the name Shewanella indica sp. nov. is proposed. The type strain is KJW27(T) ( = KCTC 23171(T)  = BCC 41031(T)  = NCIM 5388(T)).

Background: Lactobacillus plantarum is considered as a safe and effective probiotic microorganism. Among various sources of isolation, traditionally fermented foods are considered to be rich in Lactobacillus spp., which can be exploited for their probiotic attribute. Antibacterial property of L. plantarum has been demonstrated against various enteric pathogens in both in vitro and in vivo systems. This study was aimed at characterizing L. plantarum isolated from Kutajarista, an ayurvedic fermented biomedicine, and assessing its antagonistic property against a common enteropathogen Aeromonas veronii., Results: We report the isolation of L. plantarum (VR1) from Kutajarista, and efficacy of its cell free supernatant (CFS) in amelioration of cytotoxicity caused by Aeromonas veronii. On the part of probiotic attributes, VR1 was tolerant to pH 2, 0.3% bile salts and simulated gastric juice. Additionally, VR1 also exhibited adhesive property to human intestinal HT-29 cell line. Furthermore, CFS of VR1 was antibacterial to enteric pathogens like Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli, Aeromonas veronii and clinical isolates of P. aeruginosa and E. coli. Detailed study regarding the effect of VR1 CFS on A. veronii cytotoxicity showed a significant decrease in vacuole formation and detrimental cellular changes in Vero cells. On the other hand, A. veronii CFS caused disruption of tight junction proteins ZO-1 and actin in MDCK cell line, which was prevented by pre-incubation with CFS of VR1., Conclusions: This is the first study to report isolation of L. plantarum (VR1) from Kutajarista and characterisation for its probiotic attributes. Our study demonstrates the antagonistic property of VR1 to A. veronii and effect of VR1 CFS in reduction of cellular damage caused by A. veronii in both Vero and MDCK cell lines., (© 2011 Kumar et al; licensee BioMed Central Ltd.)

Two Gram-negative-staining, aerobic, non-motile, rod-shaped bacteria, designated strains FFA1(T) and FFA3(T), and belonging to the class Gammaproteobacteria were isolated from the gastrointestinal tract of adult flesh flies (Diptera: Sarcophagidae). Phylogenetic analysis of 16S rRNA gene sequence data placed these two strains within the genus Ignatzschineria with similarities of 98.6 % (FFA1(T)) and 99.35 % (FFA3(T)) to Ignatzschineria larvae L1/68(T). The level of gene sequence similarity between strains FFA1(T) and FFA3(T) was 99 %, 97.15 % and 78.1 % based on the 16S rRNA, 23S rRNA and gyrB gene sequences, respectively. Strains FFA1(T) and FFA3(T) shared 24 % DNA-DNA relatedness. DNA-DNA hybridization revealed a very low level of relatedness between the novel strains (22 % for strain FFA1(T) and 44 % for strain FFA3(T)) and I. larvae L1/68(T) genomic DNA. The respiratory quinone was Q-8 in both novel strains. The DNA G+C contents were 41.1 mol% and 40.1 mol% for strains FFA1(T) and FFA3(T), respectively. The cell membrane of both strains consisted of phosphatidylglycerol, phosphatidylethanolamine, phospholipids and aminophospholipid. The major fatty acids for both strains were C(16 : 0), summed feature 8 (C(18 : 1)ω7c and/or C(18 : 1)ω6c), CyC(19 : 0)ω8c and C(14 : 0). The results of DNA-DNA hybridization between the two new strains and I. larvae L1/68(T), in combination with phylogenetic, chemotaxonomic, biochemical and electron microscopic data, demonstrated that strains FFA1(T) and FFA3(T) represented two novel species of the genus Ignatzschineria for which the names Ignatzschineria indica sp. nov. (type strain FFA1(T) = DSM 22309(T) = KCTC 22643(T) = NCIM 5325(T)) and Ignatzschineria ureiclastica sp. nov. (type strain FFA3(T) = DSM 22310(T) = KCTC 22644(T) = NCIM 5326(T)) are proposed.

We report identification of putative innate immune related genes from a cell line of the mosquito Aedes albopictus challenged with heat-killed bacteria. Using a subtractive hybridization and sequencing approach, we analyzed a total 309 expressed sequence tags (ESTs) which clustered in 40 contigs. Thirty-five percent of genes yielded homology to known immune genes corresponding to antimicrobial peptides (AMPs), pathogen-associated molecular patterns, protease and immune signaling cascades. Interestingly, most of the genes have not been previously described from this mosquito and thus represent a class of novel immune genes. Further, 25% sequences did not match to any known species in the non-redundant databases, appear to be specific to the mosquito A. albopictus and merit further study.

This study was performed to determine whether extracellular silver nanoparticles (AgNPs) production is a genus-wide phenotype associated with all the members of genus Morganella, or only Morganella morganii RP-42 isolate is able to synthesize extracellular Ag nanoparticles. To undertake this study, all the available Morganella isolates were exposed to Ag+ ions, and the obtained nanoproducts were thoroughly analyzed using physico-chemical characterization tools such as transmission electron microscopy (TEM), UV-visible spectrophotometry (UV-vis), and X-ray diffraction (XRD) analysis. It was identified that extracellular biosynthesis of crystalline silver nanoparticles is a unique biochemical character of all the members of genus Morganella, which was found independent of environmental changes. Significantly, the inability of other closely related members of the family Enterobacteriaceae towards AgNPs synthesis strongly suggests that AgNPs synthesis in the presence of Ag+ ions is a phenotypic character that is uniquely associated with genus Morganella.

Bacillus lentus BI377, isolated from textile effluent-contaminated soil, was able to degrade 97% and 92% of Reactive Red 120 dye when 1200 and 1500 mg/l, respectively, of dye was added to nutrient broth (NB) at 35 °C within 12 h. UV-vis spectroscopy, GC-MS, FTIR and 1H NMR revealed the formation of catechol which may be further utilized by the bacterium via the TCA cycle, leading to complete mineralization. Structural analysis of metabolites in conjunction with enzyme activity studies confirmed the involvement of azoreductase, cytochrome P450 monooxygenase and other antioxidant enzymes. Decreases in total organic carbon and in biological and chemical oxygen demand suggest formation of low molecular weight metabolites that could be completely mineralized. These results suggest the potential use of B. lentus BI377 towards online treatment of textile dye effluents by using an appropriate bioreactor over a wide range of pH. This study opens-up a dependable and proficient way to use industrially viable non-pathogenic strains for biotransformation of carcinogenic dyes to ecofriendly compounds., (Copyright © 2010 Elsevier Ltd. All rights reserved.)