A novel gene from the acidophilic bacterium Leptospirillum sp. CF-1 and its role in oxidative stress and chromate tolerance

BACKGROUND: Acidophilic microorganisms like Leptospirillum sp. CF 1 thrive in environments with extremely low pH and high concentrations of dissolved heavy metals that can induce the generation of reactive oxygen species (ROS). Several hypothetical genes and proteins from Leptospirillum sp. CF 1 are known to be up regulated under oxidative stress conditions. RESULTS: In the present work, the function of hypothetical gene ABH19_09590 from Leptospirillum sp. CF 1 was studied. Heterologous expression of this gene in Escherichia coli led to an increase in the ability to grow under oxidant conditions with 5 mM K2CrO4 or 5 mM H2O2. Similarly, a significant reduction in ROS production in E. coli transformed with a plasmid carrying ABH19_09590 was observed after exposure to these oxidative stress elicitors for 30 min, compared to a strain complemented with the empty vector. A co transcriptional study using RT PCR showed that ABH19_09590 is contained in an operon, here named the "och" operon, that also contains ABH19_09585, ABH19_09595 and ABH19_09600 genes. The expression of the och operon was significantly up regulated in Leptospirillum sp. CF 1 exposed to 5 mM K2CrO4 for 15 and 30 min. Genes of this operon potentially encode a NADH:ubiquinone oxidoreductase, a CXXC motif containing protein likely involved in thiol/disulfide exchange, a hypothetical protein, and a di hydroxy acid dehydratase. A comparative genomic analysis revealed that the och operon is a characteristic genetic determinant of the Leptospirillum genus that is not present in other acidophiles. CONCLUSIONS: Altogether, these results suggest that the och operon plays a protective role against chromate and hydrogen peroxide and is an important mechanism required to face polyextremophilic conditions in acid environments.

First genome sequence of Chilean Brucella canis SCL strain provides insights on the epidemiology and virulence factors, explaining differences between geographical origins

BACKGROUND: Brucella canis is the etiological agent of canine brucellosis, a worldwide neglected zoonosis that constitutes one of the major infectious causes of infertility and reproductive failure in dogs. Although genomic information available for this pathogen has increased in recent years, here we report the first genome sequencing of a B. canis strain in Chile, and the differences in virulence genes with other B. canis strains. RESULTS: Genome assembly produced a total length of 3,289,216 bp, N50 of 95,163 and GC% of 57.27, organized in 54 contigs in chromosome I, and 21 contigs in chromosome II. The genome annotation identified a total of 1981 CDS, 3 rRNA and 36 tRNA in chromosome I, and 1113 CDS and 10 tRNA in chromosome II. There is little variation between the different strains and the SCL isolate. Phylogenetic analysis showed that the Chilean SCL strain is closely related to B. canis and B. suis strains. Small differences were found when compared to the Serbian isolate, but all strains shared the same recent common ancestor. Finally, changes in the sequence of some virulence factors showed that the SCL strain is similar to other South American B. canis strains. CONCLUSIONS: This work sequenced and characterized the complete genome of B. canis strain SCL, evidencing the complete presence of all the genes of the virB operon, and minor changes in outer membrane proteins and in the urease operon. Our data suggest that B. canis was introduced from North America and then spread throughout the South American continent.

A 14-20 kDa protein binds to the upstream region of the phtM operon involved in the synthesis of phaseolotoxin in Pseudomonas syringae pv. phaseolicola NPS3121 / Una proteína de 14-20 kDa se une a la región río arriba del operón phtM involucrado en la síntesis de faseolotoxina en Pseudomonas syringae pv. phaseolicola NPS3121

Pseudomonas syringae pv. phaseolicola is a phytopathogenic bacterium in beans that produces a phytotoxin called phaseolotoxin, in whose synthesis a group of genes that belong to the "Pht cluster" are involved. This cluster comprises 23 genes arranged in 5 transcriptional units, two monocistronic (argK, phtL) and three polycistronic (phtA, phtD, phtM) operons, whose expression is increased at 18°C, correlating with the production of phaseolotoxin by the bacterium. So far, the regulatory mechanisms involved in phaseolotoxin synthesis are poorly understood and only the requirement of low temperatures for its synthesis has been demon strated. Therefore, in this study we searched for regulatory proteins that could be involved in the phaseolotoxin synthesis, focusing on the regulation of the phtM operon. Gel shift assays showed that the promoter region of the phtM operon contains binding sites for putative regulatory proteins, which are encoded outside the Pht cluster and are independent of the GacS-GacA two-component system. Deletion assays with the promoter region of the phtM operon show that the binding site for a putative transcription factor is located within a 58 bp region. The putative transcription factor of the phtM operon has an apparent molecular mass in the 14-20 kDa range. Furthermore, the results demonstrate that the transcription factor recognizes and binds the upstream phtM region as monomer o multimer of a single polypeptide. Our findings provide new insights into the regulatory mechanisms involved in phaseolotoxin production, and suggest that the Pht cluster was integrated into the global regulatory mechanism of P. syringae pv. phaseolicola.

Biopelículas y genes icaA e icaD en estafilococos coagulasa negativos aislados de catéter endovenoso central en Unidad de Cuidados Intensivos de un Hospital Nivel III en Lima, Perú / Biofilms and icaA and icaD genes in coagulase-negative staphylococci isolated from central venous catheters from the intensive care unit of a level III hospital in Lima, Peru

Objetivo: Determinar la presencia de los genes icaA e icaD en estafilococos coagulasa negativos (SCoN) formadores de biopelículas aislados de catéteres venosos centrales (CVC) provenientes de UCI del Hospital Nacional Guillermo Almenara Irigoyen (HNGAI). Materiales y métodos: Se colectaron 151 CVC del HNGAI de Lima. Para el análisis microbiológico, se seleccionaron en forma aleatoria, 30 para el método de Maki y 121 para el método de Donlan. Se obtuvo en total, 73 aislados de SCoN, a los que se evaluó la capacidad de formar biopelículas utilizando agar rojo de Congo (ARC). A los aislados ARC positivos se les determinó la presencia de los genes icaA e icaD mediante la reacción en cadena de la polimerasa (PCR). Resultados: De 27 SCoN aislados del exterior de los CVC, 81 % (22/30) fueron ARC positivas, de los cuales 13,6 % (3/22) presentaron ambos genes; de los 46 SCoN aislados del interior de los CVC, 48 % (22/46) fueron ARC positivas, de los que 22,7 % (5/22) presentaron ambos genes. Conclusiones: 13,6 % y 22,7 % de los SCoN formadores de biopelículas y aislados del exterior e interior de CVC respectivamente, fueron portadores de ambos genes icaA e icaD.

Objective: To determine the presence of icaA and icaD genes in biofilm-forming coagulase-negative staphylococci (CoNS) isolated from central venous catheters (CVCs) from the intensive care unit of the Hospital Nacional Guillermo Almenara Irigoyen (HNGAI). Materials and methods: One hundred fifty-one (151) CVCs were collected from the HNGAI in Lima. For the microbiological analysis, 30 CVCs were randomly selected for the Maki method and 121 for the Donlan method. A total of 73 CoNS isolates were obtained and their ability to form biofilms was evaluated using Congo red agar (CRA). CRA-positive isolates showed the presence of icaA and icaD genes by polymerase chain reaction (PCR). Results: Out of the 27 CoNS isolated from the external surface of the CVCs, 81 % (22/30) were CRA positive, of which 13.6 % (3/22) presented both genes. Out of the 46 CoNS isolated from the internal surface of the CVCs, 48 % (22/46) were CRA positive, of which 22.7 % (5/22) showed both genes. Conclusions: Thirteen point six percent (13.6 %) and 22.7 % of the biofilm-forming CoNS isolated from the external and internal surfaces of the CVCs, respectively, were carriers of both icaA and icaD genes.

Evaluation of Biofilm Formation and Presence of Ica Genes in Staphylococcus epidermidis Clinical Isolates

OBJECTIVES: Biofilm formation is one of the important features of Staphylococcus epidermidis, particularly in nosocomial infections. We aimed to investigate the biofilm production by phenotypic methods and the presence of ica genes in S epidermidis. METHODS: A total of 41 S epidermidis isolates were recovered from different clinical specimens. Biofilm formation was evaluated by microtiter plate, tube method and Congo red agar method. The presence of icaA and icaD genes was investigated by PCR. Validity of methods (sensitivity and specificity), and metrics for test performance (positive/negative predictive value, and positive/negative likelihood ratio) were determined. RESULTS: By both microtiter plate and tube method, 53.6% of S epidermidis isolates were able to produce biofilm, whilst only 24.4% of isolates provided a biofilm phenotype on Congo red agar plates. icaA and icaD genes were found in 100% and 95.1% of isolates, respectively. Biofilm phenotypes accounted for 4.8% by microtiter plate assay, despite the absence of the ica gene. Congo red agar and PCR exhibited a lower sensitivity (18% and 45.5%, respectively) for identifying the biofilm phenotype in comparison to microtiter plate. CONCLUSION: The microtiter plate method remains generally a better tool to screen biofilm production in S epidermidis. In addition, the ability of S epidermidis to form biofilm is not always dependent on the presence of ica genes, highlighting the importance of ica-independent mechanisms of biofilm formation. The use of reliable methods to specifically detect biofilms can be helpful to treat the patients affected by such problematic bacteria.

Alginate overproduction and biofilm formation by psychrotolerant Pseudomonas mandelii depend on temperature in Antarctic marine sediments

Background: In recent years, Antarctica has become a key source of biotechnological resources. Native microorganisms have developed a wide range of survival strategies to adapt to the harsh Antarctic environment, including the formation of biofilms. Alginate is the principal component of the exopolysaccharide matrix in biofilms produced by Pseudomonas, and this component is highly demanded for the production of a wide variety of commercial products. There is a constant search for efficient alginate-producing organisms. Results: In this study, a novel strain of Pseudomonas mandelii isolated from Antarctica was characterized and found to overproduce alginate compared with other good alginate producers such as Pseudomonas aeruginosa and Pseudomonas fluorescens. Alginate production and expression levels of the alginate operon were highest at 4°C. It is probable that this alginate-overproducing phenotype was the result of downregulated MucA, an anti-sigma factor of AlgU. Conclusion: Because biofilm formation is an efficient bacterial strategy to overcome stressful conditions, alginate overproduction might represent the best solution for the successful adaptation of P. mandelii to the extreme temperatures of the Antarctic. Through additional research, it is possible that this novel P. mandelii strain could become an additional source for biotechnological alginate production.

Analysis of expression profile of mce operon genes (mce1, mce2, mce3 operon) in different Mycobacterium tuberculosis isolates at different growth phases.

Background & objectives: Mycobacterium tuberculosis (M. tuberculosis) has four homologous mammalian cell entry (mce) operons (mce1-4) that encode exported proteins and have a possible role in the virulence mechanism of this pathogen. The expression of mce operon is considered to be complex and not completely understood. Although expression of mce operon at different in vitro growth phases has been studied earlier, its expression in different M. tuberculosis isolates under different growth phases is not yet studied. The present preliminary study was conducted on a limited number of isolates to know the trend of expression pattern of mce operon genes in different M. tuberculosis isolates under different growth stages. Methods: In this study, we monitored the transcriptional profile of selected mce operon genes (mce1A, mce1D, mce2A, mce2D, mce3A, mce3C) in different M.tuberculosis isolates (MDR1, MDR2, and sensitive isolate) at early exponential and stationary phases using real-time quantitative PCR. Results: The expression ratio of all selected mce operon genes in all M. tuberculosis isolates was reduced at the initial phase and increased substantially at a later phase of growth. Higher expression of mce1 operon genes was found in all M. tuberculosis isolates as compared to other mce operon genes (mce2 and mce3 operons) at stationary growth phase. Interpretation & conclusions: The higher expression of mce operon genes at stationary phase (as compared to early exponential phase) suggested growth phase dependent expression of mce operon genes. This indicated that the mce operon genes might have a role in M. tuberculosis survival and adaptation on the onset of adverse condition like stationary phase. Identification of differentially expressed genes will add to our understanding of the bacilli involved in adaptation to different growth conditions.

Biomaterial implantation associted infection and related gene regulation of biofilm formation by Staphylococcus epidermidis / 国际生物医学工程杂志

Staphylococcus epidermidis is a commensal bacteria which inhabits on the surface of human skin and mucous membrane.It has been demonstrated that staphylococcus epidermidis is a major opportunistic pathogen that can cause clinical biomaterial related infection and plays an important role in biomaterial implantation associated infections.The main pathogenic factor is the formation of the bacterial biofilm on surface of medical biomaterial.The formation of bacterial biofilm can resist the defense reaction and antibiotic treatment effectively so that leading to the biomaterial implantation associated infection which is difficult to cure thoroughly,and make the infections become chronic,persistent and repetitive.Nowadays,the infections caused by Staphylococcus epidermidis have caused high clinical mortality.Staphylococcus epidermidis biofilm formation,the regulations of intercellular adhesion gene (ica) operon and accessory gene regulator (agr) gene on the formation of biofilm and their functions in clinical biomaterial related infections are reviewed in this article.

Analysis on correlation between biofilm formation and bacterial resistance in Staphylococcus epidermidis / 国际检验医学杂志

Objective To investigate the formation of biofilm in clinical isolates of Staphylococcus epidermidis ,and to analyse the correlation between biofilm formation and antibacterial resistance of Staphylococcus epidermidis .Methods A total of 62 strains of Staphylococcus epidermidis isolated from blood specimens of inpatients with bloodstream infection ,from January 2014 to February 2015 ,were collected .The biofilm formation of Staphylococcus epidermidis was detected by using the semi‐quantitative adherence as‐say and polymerase chain reaction(PCR) amplification experiment .The antibacterial susceptibility test was carried out according to K‐B method .Results The positive rate of biofilm formation detected by using the semi‐quantitative adherence assay and PCR for icaA gene were 37 .1% (23 strains) and 43 .5% (27 strains) respectively ,and there was no statistically significant difference(P>0 .05) .There were 14 positive strains detected by both methods .The resistance rates of strains producing biofilm to antibacterial a‐gents were generally higher than those of non‐producing biofilm strains ,and there were statistically significant differences in resist‐ance rates of strains to gentamicin ,penicillin ,oxacillin ,levofloxacin and cefoxitin(P<0 .05) .All bacteria were sensitive to vancomy‐cin ,linezolid and quinupristin/dalfopristin .Conclusion There is no significant difference between the two methods in detecing bio‐film formation .The resistance rates of strains producing biofilm to antibacterial agents were generally higher than those of non‐pro‐ducing biofilm strains .

Legionella dumoffii Tex-KL Mutated in an Operon Homologous to traC-traD is Defective in Epithelial Cell Invasion / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To understand the mechanism of invasion by Legionella dumoffii.</p><p><b>METHODS</b>The L. dumoffii strain Tex-KL was mutated using the Tn903 derivative, Tn903dIIlacZ. After screening 799 transposon insertion mutants, we isolated one defective mutant. We then constructed the gene-disrupted mutant, KL16, and studied its invasion of and intracellular growth in HeLa and A549 cells, and in A/J mice survival experiments. The structure of traC-traD operon was analyzed by RT-PCR.</p><p><b>RESULTS</b>The transposon insertion was in a gene homologous to Salmonella typhi traC, which is required for the assembly of F pilin into the mature F pilus structure and for conjugal DNA transmission. Results from RT-PCR suggested that the traC-traD region formed an operon. We found that when the traC gene was disrupted, invasion and intracellular growth of L. dumoffii Tex-KL were impaired in human epithelial cells. When mice were infected by intranasal inoculation with a traC deficient mutant, their survival significantly increased when compared to mice infected with the wild-type strain..</p><p><b>CONCLUSION</b>Our results indicated that the traC-traD operon is required for the invasion and intracellular growth abilities of L. dumoffii Tex-KL in epithelial cells.</p>

Diseño de péptidos basado en la secuencia análoga al represor negativo icaR de Staphylococcus sp / Design of peptides based on similar sequence of negative repressor icaR of Staphylococcus sp / Desenho de peptídeos com base na sequência similar ao repressor negativo icaR dos Staphylococcus sp

La biopelícula como un mecanismo de virulencia en Staphylococcus involucrada en infecciones intrahospitalarias es regulada por un represor negativo icaR, responsable de la transcripción completa del operón icaADBC. La búsqueda de dominios funcionales por modulación computacional de icaR permitió hallar las secuencias peptídicas con actividad biológica análoga a la proteína icaR. Mediante biología computacional se diseñaron péptidos empleando el programa de predicción AntiBP (http://www.imtech.res.in/raghava/antibp/); la síntesis química se hizo por Nα-Fmoc y se caracterizaron y purificaron tres moléculas por RP-HPLC y MALDI-TOF. Se evaluó su seguridad biológica mediante ensayo de actividad citotóxica realizada sobre macrófagos murinos de la línea J774 y la actividad hemolítica se determinó mediante el uso de glóbulos rojos. Los tres péptidos caracterizados IR1, IR2 e IR3, presentaron estructura secundaria predominantemente alfa helicoidal, alto grado de pureza y alto score antimicrobiano; además, mostraron baja toxicidad, evidenciada por la actividad citotóxica y hemolítica en las concentraciones ensayadas y en comparación con los controles usados, que permitiría su potencial uso como moléculas candidatas o principios activos con actividad análoga al represor nativo icaR, frente a la biopelícula de los Staphylococcus sp.

Staphylococcus sp. biofilm, formed as a mechanism of virulence that is involved in hospital acquired infections, is regulated by a negative repressor icaR, which is responsible for the full transcription of the operon icaADBC. This study, through functional commands by computational modulation of icaR, allowed to find peptide sequences with similar biological activity to the icaR protein. Peptides were designed by means of computational biology using the prediction program AntiBP (http://www.imtech.res.in/raghava/antibp/). The chemical synthesis of peptides was performed by Nα-Fmoc. The purification and characterization of three molecules were carried out using RP-HPLC and MALDI-TOF. Biological safety of peptides was evaluated by tests of cytotoxic activity on murine macrophage cells line J774, and their hemolytic activity was determined by using red cells. The three characterized peptides IR1, IR2 and IR3 presented a predominantly secondary alpha helical structure with a high degree of purity and high antimicrobial scores. In addition, the peptides exhibited low toxicity, proved by their low cytotoxic and hemolytic activity in the tested concentrations and in comparison to the standards used. These results allow the potential use of these peptides as candidate molecules or active principles with similar activity to the native repressor icaR against the Staphylococcus biofilm.

O biofilme formado como um mecanismo de virulência em Staphylococcus sp., que está envolvido com infecções intra-hospitalares, é regulado por um repressor negativo icaR, o qual é responsável pela plena transcrição do operão icaADBC. Por tanto, o presente estudo, avaliando a segurança biológica de moléculas, concebeu peptídeos antibiofilme semelhantes ao repressor icaR. Por meio da biologia computacional foram concebidos peptídeos usando o programa de predição AntiBP (http://www.imtech.res.in/raghava/antibp/) para identificar as sequências com uma atividade biologicamente similar à da proteína icaR. A Síntese química dos peptídeos se fez pelo Nα-Fmoc e foram caracterizadas e purificadas três moléculas por RP-HPLC e MALDI-TOF. O ensaio de atividade citotóxica foi realizado nos macrófagos murinos da linha J774 e a atividade hemolítica foi determinada por meio do uso de glóbulos vermelhos. Foram caraterizados três peptídeos IR1, IR2 e IR3, além de mostrarem uma estrutura secundaria predominantemente alfa helicoidal, com alto grau de pureza, alto score antimicrobiano e baixa toxicidade, estes podem ser postulados como moléculas candidatas ou princípios ativos com uma atividade similar ao repressor nativo icaR frente ao biofilme dos Staphylococcus sp.

Arabinose-controlled expression of htrA in Shigella flexneri through suicide plasmid-mediated homologous recombination / 军事医学

Objective To achieve arabinose-controlled expression of HtrA strain and detect the expression of HtrA protein.Methods Arabinose promoter with htrA100 was amplified from pACD-htrA vector by PCR and cloned into pGP704 vector.Then, Shigella flexneri 2a strain 301 was transferred with the recombinant plasmid pGD-htrA and an AraC-expression vector .The expressions of HtrA in whole-cell and periplasmic space were detected by Western blotting .Results The suicide plasmid-mediated homologous recombinant vector and the inducible HtrA expression strain were successfully constructed.Without arabinose,HtrA protein was hardly detected ,but in the presense of arabinose , HtrA protein could be detected in whole-cell lysate and in periplasmic space lysate by Western blot .Conclusion Homologous recombination using suicide plasmid can significantly knock down the expression of HtrA protein .After being induced with arabinose , HtrA protein can be expressed normally .

Regulation of gene expression: Cryptic β-glucoside (bgl) operon of Escherichia coli as a paradigm

Bacteria have evolved various mechanisms to extract utilizable substrates from available resources and consequently acquire fitness advantage over competitors. One of the strategies is the exploitation of cryptic cellular functions encoded by genetic systems that are silent under laboratory conditions, such as the bgl (β-glucoside) operon of E. coli. The bgl operon of Escherichia coli, involved in the uptake and utilization of aromatic β-glucosides salicin and arbutin, is maintained in a silent state in the wild type organism by the presence of structural elements in the regulatory region. This operon can be activated by mutations that disrupt these negative elements. The fact that the silent bgl operon is retained without accumulating deleterious mutations seems paradoxical from an evolutionary view point. Although this operon appears to be silent, specific physiological conditions might be able to regulate its expression and/or the operon might be carrying out function(s) apart from the utilization of aromatic β-glucosides. This is consistent with the observations that the activated operon confers a Growth Advantage in Stationary Phase (GASP) phenotype to Bgl+ cells and exerts its regulation on at least twelve downstream target genes.

Oxidative stress enhances the expression of sulfur assimilation genes: preliminary insights on the Enterococcus faecalis iron-sulfur cluster machinery regulation

The Firmicutes bacteria participate extensively in virulence and pathological processes. Enterococcus faecalis is a commensal microorganism; however, it is also a pathogenic bacterium mainly associated with nosocomial infections in immunocompromised patients. Iron-sulfur [Fe-S] clusters are inorganic prosthetic groups involved in diverse biological processes, whose in vivo formation requires several specific protein machineries. Escherichia coli is one of the most frequently studied microorganisms regarding [Fe-S] cluster biogenesis and encodes the iron-sulfur cluster and sulfur assimilation systems. In Firmicutes species, a unique operon composed of the sufCDSUB genes is responsible for [Fe-S] cluster biogenesis. The aim of this study was to investigate the potential of the E. faecalis sufCDSUB system in the [Fe-S] cluster assembly using oxidative stress and iron depletion as adverse growth conditions. Quantitative real-time polymerase chain reaction demonstrated, for the first time, that Gram-positive bacteria possess an OxyR component responsive to oxidative stress conditions, as fully described for E. coli models. Likewise, strong expression of the sufCDSUB genes was observed in low concentrations of hydrogen peroxide, indicating that the lowest concentration of oxygen free radicals inside cells, known to be highly damaging to [Fe-S] clusters, is sufficient to trigger the transcriptional machinery for prompt replacement of [Fe-S] clusters.

Revamping the role of biofilm regulating operons in device-associated Staphylococci and Pseudomonas aeruginosa.

Extensive use of indwelling devices in modern medicine has revoked higher incidence of device associated infections and most of these devices provide an ideal surface for microbial attachment to form strong biofilms. These obnoxious biofilms are responsible for persistent infections, longer hospitalization and high mortality rate. Gene regulations in bacteria play a significant role in survival, colonization and pathogenesis. Operons being a part of gene regulatory network favour cell colonization and biofilm formation in various pathogens. This review explains the functional role of various operons in biofilm expression and regulation observed in device‑associated pathogens such as Staphylococcus aureus, Staphylococcus epidermidis and Pseudomonas aeruginosa.

Glucose & sodium chloride induced biofilm production & ica operon in clinical isolates of staphylococci.

Background & objectives: All colonizing and invasive staphylococcal isolates may not produce biofilm but may turn biofilm producers in certain situations due to change in environmental factors. This study was done to test the hypothesis that non biofilm producing clinical staphylococci isolates turn biofilm producers in presence of sodium chloride (isotonic) and high concentration of glucose, irrespective of presence or absence of ica operon. Methods: Clinical isolates of 100 invasive, 50 colonizing and 50 commensal staphylococci were tested for biofilm production by microtiter plate method in different culture media (trypticase soy broth alone or supplemented with 0.9% NaCl/ 5 or 10% glucose). All isolates were tested for the presence of ica ADBC genes by PCR. Results: Biofilm production significantly increased in the presence of glucose and saline, most, when both glucose and saline were used together. All the ica positive staphylococcal isolates and some ica negative isolates turned biofilm producer in at least one of the tested culture conditions. Those remained biofilm negative in different culture conditions were all ica negative. Interpretation & conclusions: The present results showed that the use of glucose or NaCl or combination of both enhanced biofilm producing capacity of staphylococcal isolates irrespective of presence or absence of ica operon.

Bioluminescent bioreporter for assessment of arsenic contamination in water samples of India.

In the present study the most efficient R-factor controlling the ars operon was selected after screening of 39 Escherichia coli isolates by minimum inhibitory concentration test (MIC) studies from water samples of different geographical locations of India. Among all, strain isolated from Hooghly River (West Bengal) was found to have maximum tolerance towards arsenic and was further used for the development of bioreporter bacteria. Cloning of the ars regulatory element along with operator-promotor and luxCDABE from Photobacteria into expression vector has been accomplished by following recombinant DNA protocols. The bioreporter sensor system developed in this study can measure the estimated range of 0.74–60 μg of As/L and is both specific and selective for sensing bioavailable As. The constructed bacterial biosensor was further used for the determination of arsenic ion concentration in different environmental samples of India.

Comparative analysis of the genomes of Shigella dysenteriae type 2 & type 7 isolates.

Background & objectives: The four species of the genus Shigella, namely, S. dysenteriae, S. flexneri, S. boydii and S. sonnei cause a wide spectrum of illness from watery diarrhoea to severe dysentery. Genomes of these four species show great diversity. In this study, NotI, XbaI or I-CeuI restriction enzyme digested genomes of two Shigella dysenteriae isolates belonging to the serotypes 2 and 7 were extensively analyzed to find their relatedness, if any, with the whole genome sequenced strains of S. dysenteriae type 1 and S. flexneri type 2a. Methods: Pulsed-field gel electrophoresis (PFGE) technique was used to determine the diversity of Shigella genomes by rapid construction of physical maps. DNA end labelling, Southern hybridization and PCR techniques were also applied for mapping purposes. Results: The intron-coded enzyme I-CeuI cuts the bacterial genome specifically at its rrn operon. PFGE of I-CeuI digested S. dysenteriae genomes were found to carry seven rrn operons. However, I-CeuI profiles showed distinct restriction fragment polymorphism (RFLP) between the isolates as well as with the whole genome sequenced isolates. Further studies revealed that the genome sizes and I-CeuI linkage maps of the S. dysenteriae type 7 and type 2 isolates were similar to that of S. dysenteriae type 1 and S. flexneri type 2a genomes, respectively. Interpretation & conclusions: Our findings indicate that the type 7 and type 1 isolates of S. dysenteriae were probably evolved from a same precursor, while the type 2 and S. flexneri type 2a were probably evolved and diversified from a common progenitor.

Prevalence of biofilm-producing Staphylococcus epidermidis in the healthy skin of individuals in Tamil Nadu, India.

Purpose: Staphylococcus epidermidis is a major commensal bacteria. Various strains of S. epidermidis are capable of forming biofilms by attaching to several surfaces. Biofilm-forming ability of this organism is found to be associated with many hospital-acquired infections and can even impair wound healing. S. epidermidis strains producing polysaccharide-biofilms possess the intercellular adhesion (ica) operon while strains forming the protein adhesion-mediated biofilms possess the accumulation associated protein (aap) gene. We screened for biofilm-forming S. epidermidis in the skin of healthy individuals in Tamil Nadu in order to determine the risk of acquiring S. epidermidis infections in hospital settings. Materials and Methods: Skin swabs were taken from seventy two subjects residing in Chennai with healthy skin who showed no visible signs of skin lesions or allergies. S. epidermidis was isolated from 58 samples out of the 72 collected. The presence of ica operon in S. epidermidis isolates was determined by PCR and biofilm production was examined using quantitative tissue culture plate assay. Results: Majority of the samples (47/72; 65.3%) showed pure S. epidermidis growth, (14/72; 19.4%) showed pure Staphylococcus aureus growth and the remainder (11/72; 15.3%) showed mixed growth. Biofilm-forming S. epidermidis were found in the majority of samples (53/58; 91.4%) and ica operon was detected in 19 samples out of 58 (32.8%) which is a significantly higher percentage when compared to other studies conducted at different parts of the globe ( P = 0.0003). Conclusion: We inferred that ica operon and biofilm-forming S. epidermidis are common in the healthy skin of individuals in Tamil Nadu. Measures have to be taken to reduce the risk of hospital-acquired S. epidermidis infections.