Generation, annotation, and analysis of ESTs from midgut tissue of adult female Anopheles stephensi mosquitoes

<p>Abstract</p> <p>Background</p> <p>Malaria is a tropical disease caused by protozoan parasite, <it>Plasmodium</it>, which is transmitted to humans by various species of female anopheline mosquitoes. <it>Anopheles stephensi </it>is one such major malaria vector in urban parts of the Indian subcontinent. Unlike <it>Anopheles gambiae</it>, an African malaria vector, transcriptome of <it>A. stephensi </it>midgut tissue is less explored. We have therefore carried out generation, annotation, and analysis of expressed sequence tags from sugar-fed and <it>Plasmodium yoelii </it>infected blood-fed (post 24 h) adult female <it>A. stephensi </it>midgut tissue.</p> <p>Results</p> <p>We obtained 7061 and 8306 ESTs from the sugar-fed and <it>P. yoelii </it>infected mosquito midgut tissue libraries, respectively. ESTs from the combined dataset formed 1319 contigs and 2627 singlets, totaling to 3946 unique transcripts. Putative functions were assigned to 1615 (40.9%) transcripts using BLASTX against UniProtKB database. Amongst unannotated transcripts, we identified 1513 putative novel transcripts and 818 potential untranslated regions (UTRs). Statistical comparison of annotated and unannotated ESTs from the two libraries identified 119 differentially regulated genes. Out of 3946 unique transcripts, only 1387 transcripts were mapped on the <it>A. gambiae </it>genome. These also included 189 novel transcripts, which were mapped to the unannotated regions of the genome. The EST data is available as ESTDB at <url>http://mycompdb.bioinfo-portal.cdac.in/cgi-bin/est/index.cgi</url>.</p> <p>Conclusion</p> <p>3946 unique transcripts were successfully identified from the adult female <it>A. stephensi </it>midgut tissue. These data can be used for microarray development for better understanding of vector-parasite relationship and to study differences or similarities with other malaria vectors. Mapping of putative novel transcripts from <it>A. stephensi </it>on the <it>A. gambiae </it>genome proved fruitful in identification and annotation of several genes. Failure of some novel transcripts to map on the <it>A. gambiae </it>genome indicates existence of substantial genomic dissimilarities between these two potent malaria vectors.</p

Characterization of bacterial community shift in human Ulcerative Colitis patients revealed by Illumina based 16S rRNA gene amplicon sequencing

BACKGROUND: The healthy human intestine is represented by the presence of bacterial communities predominantly belonging to obligate anaerobes; however disparity and dysanaerobiosis in intestinal microflora may lead to the progression of ulcerative colitis (UC). The foremost aim of this study is to consider and compare the gut microbiota composition in patients suffering from different stages of UC. METHODS: This study represents data from the biopsy samples of six individuals suffering from UC. The samples were collected by colonoscopy and were processed immediately for isolation of DNA. Mucosal microbiota was analyzed by means of 16S rRNA gene-based Illumina high throughput sequencing. Quantitative real-time PCR (qPCR) was performed to determine total bacterial abundances. RESULTS: Analysis of 23,927 OTUs demonstrated a significant reduction of bacterial diversity consistently from phylum to species level (p < 0.05) for individuals suffering from severe stage of UC. Significant increase in abundance of unusual aerobes and facultative anaerobes, including members from the phylum Proteobacteria (p- = 0.031) was also observed. A 10 fold increase in the total bacterial count was detected in patients suffering from severe inflammatory stage (2.98 +/-0.49 E + 09/ml) when compared with patients with moderate (1.03+/-0.29 E + 08/ml) and mild (1.76 +/-0.34 E + 08/ml) stages of inflammation. CONCLUSION: The reduction of bacterial diversity with an increase in the total bacterial count indicates a shift of bacterial communities which signifies dysbiosis and dysanaerobiosis at the mucosal level for patients suffering from UC

Identity and antibiotic sensitivity pattern of the bacterial strains isolated from the PETL WWTP against 39 antibiotics.

<p>Source of strains: Strains ER1 to ER-5 and ET-1 to ET-7 were isolated from the equilibratior tank; A1R-2 to A1R-10 and A1T2 to A1T8 were isolated from aeration tank No. 1; A2R4 to A2R-8 and A2T-3 to A2T-8 were isolated from aeration tank No. 2; SR-1 to SR-14 and ST-1 to ST-8 were isolated from the settling tank; SSR-1 to SSR-6 and SST-1 to SST-8 were isolated from secondary sludge; DSR-1 to DSR-6 and DST-1 to DST-8 were isolated from dewatered sludge; OSR-1 to OSR-4 and OST-1 to OST-14 were isolated from old dried sludge.</p

Distribution of resistance among the strains from PETL WWTP against different antibiotics.

<p>Distribution of resistance among the strains from PETL WWTP against different antibiotics.</p

A Treatment Plant Receiving Waste Water from Multiple Bulk Drug Manufacturers Is a Reservoir for Highly Multi-Drug Resistant Integron-Bearing Bacteria

<div><p>The arenas and detailed mechanisms for transfer of antibiotic resistance genes between environmental bacteria and pathogens are largely unclear. Selection pressures from antibiotics in situations where environmental bacteria and human pathogens meet are expected to increase the risks for such gene transfer events. We hypothesize that waste-water treatment plants (WWTPs) serving antibiotic manufacturing industries may provide such spawning grounds, given the high bacterial densities present there together with exceptionally strong and persistent selection pressures from the antibiotic-contaminated waste. Previous analyses of effluent from an Indian industrial WWTP that processes waste from bulk drug production revealed the presence of a range of drugs, including broad spectrum antibiotics at extremely high concentrations (mg/L range). In this study, we have characterized the antibiotic resistance profiles of 93 bacterial strains sampled at different stages of the treatment process from the WWTP against 39 antibiotics belonging to 12 different classes. A large majority (86%) of the strains were resistant to 20 or more antibiotics. Although there were no classically-recognized human pathogens among the 93 isolated strains, opportunistic pathogens such as <i>Ochrobactrum intermedium, Providencia rettgeri,</i> vancomycin resistant <i>Enterococci</i> (VRE), <i>Aerococcus sp</i>. and <i>Citrobacter freundii</i> were found to be highly resistant. One of the <i>O. intermedium</i> strains (ER1) was resistant to 36 antibiotics, while <i>P. rettgeri</i> (OSR3) was resistant to 35 antibiotics. Class 1 and 2 integrons were detected in 74/93 (80%) strains each, and 88/93 (95%) strains harbored at least one type of integron. The qPCR analysis of community DNA also showed an unprecedented high prevalence of integrons, suggesting that the bacteria living under such high selective pressure have an appreciable potential for genetic exchange of resistance genes via mobile gene cassettes. The present study provides insight into the mechanisms behind and the extent of multi-drug resistance among bacteria living under an extreme antibiotic selection pressure.</p></div

Multi-drug resistance observed among 93 bacterial strains isolated from the PETL WWTP.

<p>The X-axis indicates the number of antibiotics (of 39 tested) to which the strains are resistant; the Y-axis indicates the percentage of resistant strains of 93 strains. The resistant and intermediately-resistant phenotypes are grouped together and denoted as resistant.</p

PCR-primers used in the present study.

<p>PCR-primers used in the present study.</p