scholarly journals Phylogenetic Characterization of a Novel Insect-Specific Flavivirus Detected in a Culex Pool, Collected from Assam, India

Intervirology ◽  
2015 ◽  
Vol 58 (3) ◽  
pp. 149-154 ◽  
Author(s):  
Sibnarayan Datta ◽  
Reji Gopalakrishnan ◽  
Soumya Chatterjee ◽  
Vijay Veer
Keyword(s):  
Direct Detection ◽  
Nonstructural Protein ◽  
Phylogenetic Analyses ◽  
Pcr Amplification ◽  
Specific Primers ◽  
Phylogenetic Characterization ◽  
Dna And Rna ◽  
Indian State ◽  
Mosquito Pool

Objective: We report the phylogenetic characterization of a unique flavivirus sequence detected in a wild Culex tritaeniorhynchus mosquito pool, collected from the northeast Indian state of Assam. Methods: DNA and RNA were extracted from field-collected mosquito pools. Extracts were subjected to PCR and reverse transcriptase PCR amplification using universal and type-specific primers for direct detection of flavivirus-specific viral nucleic acids. An amplified flavivirus nonstructural protein 5 (NS5) genetic region was sequenced and BLAST searched, and phylogenetic analyses performed with reference sequences retrieved from GenBank. Results: Phylogenetic analyses revealed the sequence to be related to insect-specific flaviviruses (ISFs) of the genus Flavivirus, family Flaviviridae. Despite being related to the Palm Creek virus (PCV; an ISF very recently reported from Northern Australia), the present sequence (provisionally named Assam virus) was found to be highly divergent from PCV and other ISF sequences available in GenBank. The partial NS5 sequence analysis demonstrated low nucleotide sequence identity (66-77%) with known ISFs reported from other parts of the globe. Conclusion: Findings of this study suggest the presence of a candidate novel ISF - the first to be reported from India.

scholarly journals Characterization of Microbial Community Structure in Gulf of Mexico Gas Hydrates: Comparative Analysis of DNA- and RNA-Derived Clone Libraries

2005 ◽  
Vol 71 (6) ◽  
pp. 3235-3247 ◽  
Author(s):  
Heath J. Mills ◽  
Robert J. Martinez ◽  
Sandra Story ◽  
Patricia A. Sobecky
Keyword(s):  
Phylogenetic Analysis ◽  
Microbial Community ◽  
Gulf Of Mexico ◽  
Gas Hydrate ◽  
Phylogenetic Analyses ◽  
Active Fraction ◽  
Clone Libraries ◽  
Dna And Rna ◽  
Rna And Dna

ABSTRACT The characterization of microbial assemblages within solid gas hydrate, especially those that may be physiologically active under in situ hydrate conditions, is essential to gain a better understanding of the effects and contributions of microbial activities in Gulf of Mexico (GoM) hydrate ecosystems. In this study, the composition of the Bacteria and Archaea communities was determined by 16S rRNA phylogenetic analyses of clone libraries derived from RNA and DNA extracted from sediment-entrained hydrate (SEH) and interior hydrate (IH). The hydrate was recovered from an exposed mound located in the northern GoM continental slope with a hydrate chipper designed for use on the manned-submersible Johnson Sea Link (water depth, 550 m). Previous geochemical analyses indicated that there was increased metabolic activity in the SEH compared to the IH layer (B. N. Orcutt, A. Boetius, S. K. Lugo, I. R. Macdonald, V. A. Samarkin, and S. Joye, Chem. Geol. 205:239-251). Phylogenetic analysis of RNA- and DNA-derived clones indicated that there was greater diversity in the SEH libraries than in the IH libraries. A majority of the clones obtained from the metabolically active fraction of the microbial community were most closely related to putative sulfate-reducing bacteria and anaerobic methane-oxidizing archaea. Several novel bacterial and archaeal phylotypes for which there were no previously identified closely related cultured isolates were detected in the RNA- and DNA-derived clone libraries. This study was the first phylogenetic analysis of the metabolically active fraction of the microbial community extant in the distinct SEH and IH layers of GoM gas hydrate.

scholarly journals First Report of Anthracnose Caused by Colletotrichum gloeosporioides in Pumpkin in Trinidad

Plant Disease ◽  
2010 ◽  
Vol 94 (8) ◽  
pp. 1062-1062
Author(s):  
S. N. Rampersad
Keyword(s):  
Colletotrichum Gloeosporioides ◽  
Phylogenetic Analyses ◽  
Pcr Amplification ◽  
Positive Control ◽  
Universal Primers ◽  
Conidial Suspension ◽  
Specific Primers ◽  
First Report ◽  
Main Production ◽  
Species Specific

In Trinidad, pumpkin (Cucurbita pepo L. and C. moschata L.) is extensively grown for local and international export markets. In November 2008, symptoms of foliar chlorosis and necrosis were observed in 15 commercial pumpkin fields located in the main production areas of St. George East, Caroni, Victoria, and St. Patrick counties. Severely infected plants were unable to support fruit maturation, which resulted in yield loss. The pathogen was isolated from surface-sterilized tissues of symptomatic plants. Colonies on potato dextrose agar (PDA) were white to cream with gray spore masses in the center. Conidia were hyaline, cylindrical with rounded ends, aseptate, and measured 12.5 to 16.5 μm × 3.5 to 5.0 μm. PCR amplification was carried out with ITS4/5 universal primers (4) and species-specific primers, CgInt/ITS4 (1), using a positive control of Colletotrichum gloeosporioides (courtesy of D. Perez-Brito). Species-specific primers generated a single amplicon, ~450 bp long, which corresponded with the positive control. The ITS1 region (1) of pumpkin isolates (GenBank No. GU320190) was 100% identical to cognate sequences of C. gloeosporioides isolates (GenBank Nos. AY841136 and FJ624257). Phylogenetic analyses (MEGA 4 – Molecular Evolutionary Genetic Analysis Software version 4 for Windows) using the neighbor-joining (NJ) algorithm placed the pumpkin isolates in a well-supported cluster (>90% bootstrap value based on 1,000 replicates) with other C. gloeosporioides isolates. The tree was rooted with C. crassipes (GenBank No. AJ536230). The pathogen was similar to C. gloeosporioides (Penz.) Penz. & Sacc. (3). In pathogenicity tests, six plants (cv. Jamaican squash) for each of five isolates were spray inoculated to runoff with a conidial suspension (1.0 × 106 conidia/ml). Negative controls were sprayed with sterile distilled water. In repeated tests, plants were symptomatic of infection 7 days postinoculation. There were no symptoms on control plants. Koch's postulates were fulfilled with the reisolation of the pathogen from symptomatic leaf tissues. Anthracnose is a serious threat to cucurbit production; however, infection is not common in pumpkin and squash (2). To my knowledge, this is the first report of C. gloeosporioides causing widespread anthracnose infection in pumpkin in Trinidad. References: (1) A. E. Brown et al. Phytopathology 86:523, 1996. (2) G. Kelly. Acta Hortic. (ISHS) 731:479, 2007. (3) B. C. Sutton. Page 1 in: Colletotrichum: Biology, Pathology and Control. CAB International. Wallingford, UK, 1992. (4) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990.

scholarly journals CHARACTERIZATION OF BACTERIAL ISOLATES FOR SUSTAINABLE RICE BLAST CONTROL

2020 ◽  
Vol 33 (3) ◽  
pp. 702-712
Author(s):  
BÁRBARA ESTEVAM DE MELO MARTINS ◽  
AMANDA ABDALLAH CHAIBUB ◽  
MARCIO VINICIUS DE CARVALHO BARROS CORTÊS ◽  
VALÁCIA LEMES DA SILVA LOBO ◽  
MARTA CRISTINA CORSI DE FILIPPI
Keyword(s):  
Rice Blast ◽  
Pcr Amplification ◽  
Morphological Characterization ◽  
Bacterial Isolates ◽  
Specific Primers ◽  
Leaf Blast ◽  
Fluorescent Pigment ◽  
Gram Staining ◽  
Bacillus Spp

ABSTRACT Rice blast (Magnaporthe oryzae) limits rice (Oryza sativa) grain yields worldwide. The objective of this investigation was to morphologically, biochemically, and molecularly characterize six bacterial isolates, BRM 32109, BRM 32110, BRM 32111, BRM 32112, BRM 32113, and BRM 32114, and to determine their potential as antagonists to M. oryzae. Morphological characterization was based on colony formation and color, Gram staining, and fluorescent pigment production. Biochemical studies were based on cellulase, chitinase, phosphatase, indoleacetic acid, and siderophore production, as well as biofilm formation. The molecular identification used specific primers for PCR amplification of the 16S rRNA region, followed by sequencing. The antagonism studies involved three experiments, which had randomized designs. Two of them were conducted in laboratory conditions, pairing bacterial colonies and M. oryzae, using bacterial filtrates, and the third was conducted in greenhouse conditions. BRM 32111 and BRM 32112 were identified as Pseudomonas sp., BRM 32113 as Burkholderia sp., BRM 32114 as Serratia sp., and BRM 32110 and BRM 32109 as Bacillus spp. BRM 32112, BRM 32111, and BRM 32113 inhibited the colony of M. oryzae by 68%, 65%, and 48%, respectively. The bacterial suspensions of the BRM 32111, BRM 32112, and BRM 3212 filtrates suppressed leaf blast by 81.0, 79.2, and 66.3%, respectively. BRM 32111 and BRM 32112 were determined to be antagonists of M. oryzae and were found to solubilize phosphate, produce siderophores and cellulose, form biofilms, and suppress leaf blast. These isolates should be further investigated as potential biological control agents for leaf blast control.

scholarly journals Characterization of a Novel Spirochete Associated with the Hydrothermal Vent Polychaete Annelid, Alvinella pompejana

2001 ◽  
Vol 67 (1) ◽  
pp. 110-117 ◽  
Author(s):  
Barbara J. Campbell ◽  
S. Craig Cary
Keyword(s):  
High Temperature ◽  
Hydrothermal Vent ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Pcr Amplification ◽  
Dorsal Surface ◽  
Specific Primers ◽  
Gradient Gel Electrophoresis ◽  
Polychaetous Annelid ◽  
Alvinella Pompejana

ABSTRACT A highly integrated, morphologically diverse bacterial community is associated with the dorsal surface of Alvinella pompejana, a polychaetous annelid that inhabits active high-temperature deep-sea hydrothermal vent sites along the East Pacific Rise (EPR). Analysis of a previously prepared bacterial 16S ribosomal DNA (rDNA) library identified a spirochete most closely related to an endosymbiont of the oligochete Olavius loisae. This spirochete phylotype (spirochete A) comprised only 2.2% of the 16S rDNA clone library but appeared to be much more dominant when the same sample was analyzed by denaturing gradient gel electrophoresis (DGGE) and the terminal restriction fragment length polymorphism procedure (12 to 18%). PCR amplification of the community with spirochete-specific primers used in conjunction with DGGE analysis identified two spirochete phylotypes. The first spirochete was identical to spirochete A but was present in only one A. pompejana specimen. The second spirochete (spirochete B) was 84.5% similar to spirochete A and, more interestingly, was present in the epibiont communities of all of theA. pompejana specimens sampled throughout the geographic range of the worm (13°N to 32°S along the EPR). The sequence variation of the spirochete B phylotype was less than 3% for the range of A. pompejana specimens tested, suggesting that a single spirochete species was present in the A. pompejanaepibiotic community. Additional analysis of the environments surrounding the worm revealed that spirochetes are a ubiquitous component of high-temperature vents and may play an important role in this unique ecosystem.

scholarly journals Histopathology and Molecular Characterization of Microsporidian Parasites in Cultured Shrimp Penaeus Monodon Fabricius 1798 of Bangladesh

2019 ◽  
Vol 45 (2) ◽  
pp. 187-196 ◽  
Author(s):  
Md Munjur Hossain ◽  
Nusrat Jahan Punom ◽  
Md Mostavi Enan Eshik ◽  
Mst Khadiza Begum ◽  
Md Aminul Islam Bhuiyan ◽  
...  
Keyword(s):  
Penaeus Monodon ◽  
Pcr Amplification ◽  
Histological Section ◽  
Rrna Gene ◽  
Microsporidian Parasite ◽  
Specific Primers ◽  
West Region ◽  
Field Samples ◽  
Enterocytozoon Hepatopenaei

Black tiger shrimp (Penaeus monodon Fabricius 1798) cultured in Bangladesh was investigated for the presence of microsporidian parasite. Histological section of hepatopancreas showed a large number of microsporidian spores under light microscopy. Spores under scanning electron microscope appeared oval shapes. Histology of infected shrimps showed severe degeneration of hepatopancreatic tubules. Early and late stage of microsporidian parasites in hepatopancreatic tubules were also observed. DNA extracted from the hepatopancreas of shrimps were subjected to PCR amplification using primers targeting microsporidian SSU rRNA gene. The PCR amplified an expected product of ~328 bp and the sequences showed 81 - 82% identity with the Paranucleospora theridion reported from western Norway in 2008. Further screening of field samples was carried out using EHP-specific primers. DNA extracted from ten hepatopancreas samples of P. monodon were tested and none found to be positive for EHP (Enterocytozoon hepatopenaei). This is the first report for the identification of microsporidian parasites in cultured shrimp along the south-west region of Bangladesh. Asiat. Soc. Bangladesh, Sci. 45(2): 187-196, December 2019

scholarly journals The ompA Gene in Chlamydia trachomatis Differs in Phylogeny and Rate of Evolution from Other Regions of the Genome

2006 ◽  
Vol 74 (1) ◽  
pp. 578-585 ◽  
Author(s):  
Brian W. Brunelle ◽  
George F. Sensabaugh
Keyword(s):  
Chlamydia Trachomatis ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Sequence Data ◽  
Phylogenetic Analyses ◽  
Housekeeping Genes ◽  
Phylogenetic Characterization ◽  
Tissue Tropisms

ABSTRACT Strains of Chlamydia trachomatis are classified into serovars based on nucleotide sequence differences in ompA, the gene that encodes the major outer membrane protein. Phylogenetic characterization of strains based on ompA, however, results in serovar groupings that are inconsistent with the distinguishing features of C. trachomatis pathobiology, e.g., tissue tropisms and disease presentation. We have compared nucleotide sequences at multiple sites distributed around the chlamydial genome from 18 strains representing 16 serovars; sampled regions included genes encoding housekeeping enzymes (totaling 2,073 bp), intergenic noncoding segments (1,612 bp), and a gene encoding a second outer membrane protein (porB; 1,023 bp), with the ompA sequence (1,194 bp) used for reference. These comparative analyses revealed substantial variation in nucleotide substitution patterns among the sampled regions, with average pairwise sequence differences ranging from 0.15% for the housekeeping genes to 12.1% for ompA. Phylogenetic characterization of the sampled genomic sequences yielded a strongly supported tree that divides the strains into groupings consistent with C. trachomatis biology and which has a topology quite distinct from the ompA tree. This phylogenetic incongruity can be accounted for by recombination of the ompA gene between different genomic backgrounds. We found, however, no evidence of recombination within or between any of the sampled regions around the C. trachomatis genome apart from ompA. Parallel analysis of published sequence data on four members of the pmp gene family are consistent with the phylogenetic analyses reported here.

scholarly journals Genetic and phylogenetic characterization of genome segments 2 and 6 of bluetongue virus isolates in Japan from 1985 to 2008

2012 ◽  
Vol 93 (7) ◽  
pp. 1465-1473 ◽  
Author(s):  
Hiroaki Shirafuji ◽  
Tohru Yanase ◽  
Tomoko Kato ◽  
Makoto Yamakawa
Keyword(s):  
Bluetongue Virus ◽  
Phylogenetic Analyses ◽  
Control Measures ◽  
Asia Pacific ◽  
Segment Length ◽  
Genetic Characteristics ◽  
Phylogenetic Characterization ◽  
Partial Correlations ◽  
Different Origins

This study conducted genetic and phylogenetic analyses of genome segments 2 and 6 (Seg-2 and Seg-6), which encode serotype-specific structural proteins of the outer capsid, of bluetongue virus (BTV) isolated in Japan from 1985 to 2008. The Japanese strains of BTV were clearly sorted into six groups by several genetic characteristics of Seg-2, including segment length, ORF length and 5′- and 3′-terminal sequences, and were identified as serotypes 2, 3, 9, 12, 16 and 21 by phylogenetic comparisons with Seg-2 of reference and field strains of serotypes 1–24. In contrast, phylogenetic comparisons of Seg-6 also revealed some variations among the Japanese strains and partial correlations of the serotypes between the Japanese strains and the reference or field strains. Thus, the results revealed that at least six serotypes of BTV were isolated in Japan and that there were some variations in the genetic and phylogenetic characteristics of Seg-2 and Seg-6 among the Japanese strains, suggesting that BTV of several different origins has appeared sporadically in Japan. These data will be beneficial for understanding BTV epidemiology and taking better control measures against bluetongue in Japan and its neighbouring countries in the Asia-Pacific region.

Morphological, physiological, molecular and phylogenetic characterization of new environmental isolates of Acanthamoeba spp. from the region of Bratislava, Slovakia

Biologia ◽  
2010 ◽  
Vol 65 (1) ◽  
Author(s):  
Viera Nagyová ◽  
Arpád Nagy ◽  
Štefan Janeček ◽  
Jozef Timko
Keyword(s):  
Pcr Amplification ◽  
Temperature Tolerance ◽  
Environmental Isolates ◽  
Environmental Distribution ◽  
Pathogenic Potential ◽  
Phylogenetic Characterization ◽  
Group I ◽  
Genotype Identification

AbstractProtozoa of the genus Acanthamoeba are organisms that can be generally found in the environment. The focus of this study is the detection of the presence of Acanthamoeba in different water sources and samples taken from airconditioning units. The identification of Acanthamoeba isolates was based on the morphology of cysts and trophozoites as well as PCR amplification with a genus specific primer pair JDP1 and JDP2. Growth characteristics and temperature tolerance were monitored. The pathogenic potential was tested in vitro on Vero cell cultures. Genotype identification was based on the sequencing of the GTSA.B1 PCR amplimer of 18S ribosomal DNA. The data obtained revealed that the isolates belong to T3 and T4 genotypes. One T3 and one T4 isolate contain a group I intron. The 933 base pair intron found in a genotype T4 isolate is considerably larger compared to formerly described introns of Acanthamoeba griffini (genotype T3) and A. Lenticulata (genotype T5). This is the first report detailing the environmental distribution of the Acanthamoeba genotypes in the region of Bratislava, Slovakia.

scholarly journals Typing of Intimin Genes in Human and Animal Enterohemorrhagic and Enteropathogenic Escherichia coli: Characterization of a New Intimin Variant

2000 ◽  
Vol 68 (1) ◽  
pp. 64-71 ◽  
Author(s):  
E. Oswald ◽  
H. Schmidt ◽  
S. Morabito ◽  
H. Karch ◽  
O. Marchès ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Animal Species ◽  
Pcr Amplification ◽  
Binding Activity ◽  
Cell Binding ◽  
Specific Primers ◽  
E Coli ◽  
Specific Manner ◽  
Tight Association

ABSTRACT Enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC) produce the characteristic “attaching and effacing” (A/E) lesion of the brush border. Intimin, an outer membrane protein encoded by eae, is responsible for the tight association of both pathogens with the host cell. Severaleae have been cloned from different EPEC and EHEC strains isolated from humans and animals. These sequences are conserved in the N-terminal region but highly variable in the last C-terminal 280 amino acids (aa), where the cell binding activity is localized. Based on these considerations, we developed a panel of specific primers to investigate the eae heterogeneity of the variable 3′ region by using PCR amplification. We then investigated the distribution of the known intimin types in a large collection of EPEC and EHEC strains isolated from humans and different animal species. The existence of a yet-unknown family of intimin was suspected because several EHEC strains, isolated from human and cattle, did not react with any of the specific primer pairs, although these strains were eaepositive when primers amplifying the conserved 5′ end were used. We then cloned and sequenced the eae present in one of these strains (EHEC of serotype O103:H2) and subsequently designed a PCR primer that recognizes in a specific manner the variable 3′ region of this new intimin type. This intimin, referred to as “ɛ,” was present in human and bovine EHEC strains of serogroups O8, O11, O45, O103, O121, and O165. Intimin ɛ is the largest intimin cloned to date (948 aa) and shares the greatest overall sequence identity with intimin β, although analysis of the last C-terminal 280 aa suggests a greater similarity with intimins α and γ.

scholarly journals Physiological, Ecological, and Phylogenetic Characterization of Stappia, a Marine CO-Oxidizing Bacterial Genus

2006 ◽  
Vol 73 (4) ◽  
pp. 1266-1276 ◽  
Author(s):  
Carolyn F. Weber ◽  
Gary M. King
Keyword(s):  
Phylogenetic Analyses ◽  
Large Subunit ◽  
Membrane Lipid ◽  
Rrna Gene ◽  
16S Rrna Gene Sequences ◽  
Bisphosphate Carboxylase ◽  
Phylogenetic Characterization ◽  
Metabolic Versatility ◽  
Wide Range

ABSTRACT Bacteria play a major role in marine CO cycling, yet very little is known about the microbes involved. Thirteen CO-oxidizing Stappia isolates obtained from existing cultures, macroalgae, or surf samples representing geographically and ecologically diverse habitats were characterized using biochemical, physiological, and phylogenetic approaches. All isolates were aerobic chemoorganotrophs that oxidized CO at elevated (1,000 ppm) and ambient-to-subambient concentrations (<0.3 ppm). All contained the form I (OMP) coxL gene for aerobic CO dehydrogenase and also the form II (BMS) putative coxL gene. In addition, some strains possessed cbbL, the large subunit gene for ribulose-1,5-bisphosphate carboxylase/oxygenase, suggesting the possibility of lithotrophic or mixotrophic metabolism. All isolates used a wide range of sugars, organic acids, amino acids, and aromatics for growth and grew at salinities from 5 to 45 ppt. All but one isolate denitrified or respired nitrate. Phylogenetic analyses based on 16S rRNA gene sequences indicated that several isolates could not be distinguished from Stappia aggregata and contributed to a widely distributed species complex. Four isolates (of strains GA15, HI, MIO, and M4) were phylogenetically distinct from validly described Stappia species and closely related genera (e.g., Ahrensia, Pannonibacter, Pseudovibrio, and Roseibium). Substrate utilization profiles, enzymatic activity, and membrane lipid composition further distinguished these isolates and supported their designations as new Stappia species. The observed metabolic versatility of Stappia likely accounts for its cosmopolitan distribution and its ability to contribute to CO cycling as well as other important biogeochemical cycles.

Sign in / Sign up

Export Citation Format

Share Document