Strain GA Research Articles

Phylogenetic and pathogenic characterization of Mauginiella scaettae as the causal agent of date palm (Phoenix dactylifera L.) inflorescence rot in southeast of Algeria

Inflorescence rot is a devastating disease of date palm (Phoenix dactylifera L.) but has not been extensively characterised. Mauginiella scaettae Cav. 1925 (Pleosporales; Ascomycota), causal agent of this disease, was isolated from infected male and female inflorescences of date palm from different oases in the southeast of Algeria: Ouargla (strains OU1, OU2, OU3), Tougourt (strains TO1, TO2), Hadjira (strain HA), Oued (strains OE1, OE2), Ghardaia (strain GA) and Biskra (strain BI). Each strain of M. scaettae was used to infect healthy male inflorescence of date palms to satisfy Koch's postulates but follow on assessments were based on GA. Phylogenetic reconstruction using the rRNA sequence data (ITS and LSU regions) found no intraspecific differentiation of strains and confirmed the placement of M. scaettae within the family Phaeosphaeriaceae. Fourier Transform Infrared spectroscopy (FTIR) metabolite fingerprinting discriminated between fungal strains originating from male and female inflorescences. Scanning electron microscopy (SEM) characterisation of the infection processes for M. scaettae in both male and female inflorescences suggested a preference for stomatal entry. Our characterisation provides new insights into this inflorescence rot to allow better detection and management of the disease.

A Genome-Centric Approach Reveals a Novel Glycosyltransferase from the GA A07 Strain of Bacillus thuringiensis Responsible for Catalyzing 15-O-Glycosylation of Ganoderic Acid A.

Strain GA A07 was identified as an intestinal Bacillus bacterium of zebrafish, which has high efficiency to biotransform the triterpenoid, ganoderic acid A (GAA), into GAA-15-O-β-glucoside. To date, only two known enzymes (BsUGT398 and BsUGT489) of Bacillus subtilis ATCC 6633 strain can biotransform GAA. It is thus worthwhile to identify the responsible genes of strain GA A07 by whole genome sequencing. A complete genome of strain GA A07 was successfully assembled. A phylogenomic analysis revealed the species of the GA A07 strain to be Bacillus thuringiensis. Forty glycosyltransferase (GT) family genes were identified from the complete genome, among which three genes (FQZ25_16345, FQZ25_19840, and FQZ25_19010) were closely related to BsUGT398 and BsUGT489. Two of the three candidate genes, FQZ25_16345 and FQZ25_19010, were successfully cloned and expressed in a soluble form in Escherichia coli, and the corresponding proteins, BtGT_16345 and BtGT_19010, were purified for a biotransformation activity assay. An ultra-performance liquid chromatographic analysis further confirmed that only the purified BtGT_16345 had the key biotransformation activity of catalyzing GAA into GAA-15-O-β-glucoside. The suitable conditions for this enzyme activity were pH 7.5, 10 mM of magnesium ions, and 30 °C. In addition, BtGT_16345 showed glycosylation activity toward seven flavonoids (apigenein, quercetein, naringenein, resveratrol, genistein, daidzein, and 8-hydroxydaidzein) and two triterpenoids (GAA and antcin K). A kinetic study showed that the catalytic efficiency (kcat/KM) of BtGT_16345 was not significantly different compared with either BsUGT398 or BsUGT489. In short, this study identified BtGT_16345 from B. thuringiensis GA A07 is the catalytic enzyme responsible for the 15-O-glycosylation of GAA and it was also regioselective toward triterpenoid substrates.

Endophytic Bacillus and Pseudomonas spp. Modulate Apple Shoot Growth, Cellular Redox Balance, and Protein Expression Under in Vitro Conditions.

Interactions between host plants and endophytic microorganisms play an important role in plant responses to pathogens and environmental stresses and have potential applications for plant stress management under in vitro conditions. We assessed the effect of endophytic bacteria on the growth and proliferation of domestic apple cv. Gala shoots in vitro. Further, a model apple cell suspension system was used to examine molecular events and protein expression patterns at an early stage of plant–endophyte interaction. Among the seven strains used in the study, Bacillus spp. strains Da_1, Da_4, and Da_5 and the Pseudomonas fluorescens strain Ga_1 promoted shoot growth and auxiliary shoot proliferation. In contrast, Bacillus sp. strain Oa_4, P. fluorescens strain Ga_3 and P. orientalis strain G_12 inhibited shoot development. In the cell suspension, the effects of the association between endophytic bacteria and plant cells were specific to each strain. Modulation of the cellular redox balance was monitored in the apple cells using a 2′,7′-dichlorodihydrofluorescein diacetate (H2DCFDA) probe, and strain-specific effects were observed that correlated with the in vitro shoot development results. Proteomic analysis revealed differences in protein expressions in apple cells co-cultivated with different Bacillus spp. strains that had contrasting effects on cellular redox balance and shoot development. The Bacillus sp. strain Da_4, which enhanced shoot development and oxidation of H2DCFDA, induced differential expression of proteins that are mainly involved in the defense response and regulation of oxidative stress. Meanwhile, treatment with Bacillus sp. strain Oa_4 led to strong upregulation of PLAT1, HSC70-1 and several other proteins involved in protein metabolism and cell development. Taken together, the results suggest that different cell signaling and response events at the early stage of the plant–endophyte interaction may be important for strain-dependent regulation of cellular redox balance and development of shoot phenotype.

Synergy of carbon and nitrogen removal of a co-culture of two aerobic denitrifying bacterial strains, Acinetobacter sp. GA and Pseudomonas sp. GP.

Two newly isolated aerobic denitrifying bacterial strains (Acinetobacter sp. GA and Pseudomonas sp. GP) were co-cultured to investigate the synergy of carbon and nitrogen removal of different functional bacteria. The co-culture showed higher efficiency for removing total organic carbon (TOC) and total dissolved nitrogen (TDN) than strain GA or GP cultured separately. The inoculation ratio of 50 (strain GA/GP) was advantageous for the TOC and TDN removal efficiencies of the co-culture. The sequential co-culture tests showed that strain GP being inoculated after strain GA was inoculated for 36 h increased the TDN removal efficiency from 53.3% to 86.8%. This finding indicated that the activity of strain GA was important in the co-culture system, and the sequential co-culture could be advantageous to the synergistic effect of strains GA and GP. The co-culture of different functional bacteria can be an alternative method for improving the performance of aerobic denitrifying microorganisms for pollutant removal.

Repurposing of Anticancer Drugs for the Treatment of Bacterial Infections.

Despite the fact that bacterial infections are one of the leading causes of death worldwide and that mortality rates are increasing at alarming rates, no new antibiotics have been produced by the pharmaceutical industry in more than a decade. The situation is so dire that the World Health Organization warned that we may enter a "post-antibiotic era" within this century; accordingly, bacteria resistant against all known antibiotics are becoming common and already producing untreatable infections. Although several novel approaches to combat bacterial infections have been proposed, they have yet to be implemented in clinical practice. Hence, we propose that a more plausible and faster approach is the utilization of drugs originally developed for other purposes besides antimicrobial activity. Among these are some anticancer molecules proven effective in vitro for eliminating recalcitrant, multidrug tolerant bacteria; some of which also protect animals from infections and recently are undergoing clinical trials. In this review, we highlight the similarities between cancer cells/tumors and bacterial infections, and present evidence that supports the utilization of some anticancer drugs, including 5-fluorouracil (5-FU), gallium (Ga) compounds, and mitomycin C, as antibacterials. Each of these drugs has some promising properties such as broad activity (all three compounds), dual antibiotic and antivirulence properties (5-FU), efficacy against multidrug resistant strains (Ga), and the ability to kill metabolically dormant persister cells which cause chronic infections (mitomycin C).

Characterization of a Gallid herpesvirus 2 strain with novel reticuloendotheliosis virus long terminal repeat inserts.

A bacterial artificial chromosome clone, designated LCY, was constructed from a Gallid herpesvirus 2 (GaHV-2) isolate from a GaHV-2 and reticuloendotheliosis virus co-infected clinical sample. The LCY GaHV-2 insert was sequenced and found to consist of 175,319 nucleotides. LCY GaHV-2 open reading frames (ORFs) had a high sequence identity to those of reference strains. The major difference was that two REV long terminal repeats (LTRs), in the same direction, were inserted at the internal repeat short (IRs)/unique short (Us) and Us/terminal repeat short (TRs) junctions. In addition, the a-like sequence and UL36 were different from other strains. Phylogenetic analysis revealed that LCY was closely related to pandemic strains in China. A pathogenicity study and a vaccination-challenge test were performed on LCY and the reference strain, GA. The results showed that LCY induced gross Marek's disease (MD) lesions and mortality in 71.4 and 7.1% of chickens, respectively, which are lower rates than those observed for the reference strain GA (85.7 and 35.7%). The commercially available CVI988 vaccine provided complete protection against LCY and GA (100%). These results showed that the isolate exhibited lower pathogenicity in SPF chickens. This study revealed that a novel pattern of LTR inserts was found in the strain LCY and that the strain was of low virulence. The present work expands the available genetic information for GaHV-2 and will be useful for the control of MD in China.

Molecular and pathogenicity characterization of Gallid herpesvirus 2 newly isolated in China from 2009 to 2013

During the course of our continuous surveillance of Gallid herpesvirus 2 (GaHV-2), 44 isolates were obtained from GaHV-2-positive chickens of different flocks in China from 2009 to 2013. The meq gene, considered as a major GaHV-2 oncogene, was sequenced and was found to contain an open reading frame of 1020 nucleotides encoding a 339 amino acid (aa) polypeptide in all isolates. Compared with the GaHV-2 GA strain, the meq genes in 15.9 % (7/44) of the isolates analyzed in this study contained an aa substitution mutation at position 88 (A to T) of which is the first report. The main characteristics of Chinese GaHV-2 isolates meq genes included the substitutions K77E, D80Y, V115A, T139A, P176R, and P217A, and the aa substitution frequency at positions 139 and 176 showed an increase. To test the pathogenicity of the isolates, a pathogenicity study and a vaccination-challenge test were performed on three selected isolates (ZY/1203, WC/1203, and WC/1110) and reference strain GA. The results showed that the three isolates induced gross Marek's disease (MD) lesions in 95.0-100 % cases, which was a higher rate than that obtained for strain GA (82.4 %). Three isolates induced mortality in 10-21.1 % of specific-pathogen-free chickens, which was similar to results with strain GA (23.5 %). The commercially available CVI988 vaccine induced lower protective indices (PIs) against ZY/1203 (82.4) and WC/1110 (83.3) as compared to those against WC/1203 (100) and GA (100). These results showed an evolving trend in the meq genes of the isolates; three isolates exhibited higher morbidity as compared to the reference strain and the vaccine induced lower PIs against two isolates as compared to that against the reference strain.

Easy axis reorientation and magneto-crystalline anisotropic resistance of tensile strained (Ga,Mn)As films

We present a study of magnetic anisotropy by using magneto-transport and direct magnetization measurements on tensile strained (Ga,Mn)As films. The magnetic easy axis of the films is in-plane at low temperatures, while the easy axis flips to out-of-plane when temperature is raised or hole concentration is increased. This easy axis reorientation is explained qualitatively in a simple physical picture by Zener’s p–d model. In addition, the magneto-crystalline anisotropic resistance was also investigated experimentally and theoretically based on the single magnetic domain model. The dependence of sheet resistance on the angle between the magnetic field and [1 0 0] direction was measured. It is found that the magnetization vector M in the single-domain state deviates from the external magnetic field H direction at low magnetic field, while for high magnetic field, M continuously moves following the field direction, which leads to different resistivity function behaviors.

Theoretical analysis of the influence of magnetic domain walls on longitudinal and transverse magnetoresistance in tensile strained (Ga,Mn)As epilayers

We present a theoretical analysis of recent experimental measurements of magnetoresistance in ${\mathrm{Ga}}_{1\ensuremath{-}x}{\mathrm{Mn}}_{x}\mathrm{As}$ epilayers with perpendicular magnetic anisotropy. The model reproduces the field-antisymmetric anomalies observed in the longitudinal magnetoresistance in the planar geometry (magnetic field in the epilayer plane and parallel to the current density), as well as the unusual shape of the accompanying transverse magnetoresistance. The magnetoresistance characteristics are attributed to circulating currents created by the presence of magnetic domain walls.

Isolation and Characterization of Staphylococci from External Auditory Meatus of Dogs with or without Otitis Externa with Special Reference to Staphylococcus schleiferi subsp. coagulans Isolates

Staphylococci were isolated from the external auditory meatus in 14 (48.3%) of 29 dogs affected with otitis externa (OE dogs) and 28 (68.3%) of 41 dogs without OE (non-OE dogs). Twenty-two OE isolates were identified as belonging to 12 species, and 42 non-OE isolates were identified as belonging to 13 species. The predominant species found in both OE and non-OE isolates were S. intermedius, and S. epidermidis. Thirty-eight (59.4%) of 64 isolates were resistant to one or more of the 17 antimicrobial agents tested. Resistance to PCG and ABPC was most frequent. S. schleiferi subsp. coagulans, a recent etiologic agent of canine OE, was isolated from OE and non-OE dogs. All of the 5 S. schleiferi subsp. coagulans isolates showed typical characteristics. No clear difference in the extracellular enzyme or toxin profiles, nor in the PFGE patterns, was demonstrated between the OE and non-OE isolates of S. schleiferi subsp. coagulans. A new PCR primer set specific for 16S rDNA was designed to identify strains of S. schleiferi subsp. coagulans. The amplified fragment was detected in all of the 5 isolates as well as the type strain GA 211 (=JCM 7470) and a reference strain GA 11, but was not detected in any strains of the related species, S. aureus, S. intermedius and S. hyicus. The PCR may allow a simple, rapid and precise identification of S. schleiferi subsp. coagulans, in addition to the standard tube test for free coagulase.

Characterization of an infectious cDNA copy of the genome of a naturally occurring, avirulent coxsackievirus B3 clinical isolate

Group B coxsackieviruses (CVB) cause numerous diseases, including myocarditis, pancreatitis, aseptic meningitis and possibly type 1 diabetes. To date, infectious cDNA copies of CVB type 3 (CVB3) genomes have all been derived from pathogenic virus strains. An infectious cDNA copy of the well-characterized, non-pathogenic CVB3 strain GA genome was cloned in order to facilitate mapping of the CVB genes that influence expression of a virulence phenotype. Comparison of the sequence of the parental CVB3/GA population, derived by direct RT-PCR-mediated sequence analysis, to that of the infectious CVB3/GA progeny genome demonstrated that an authentic copy was cloned; numerous differences were observed in coding and non-coding sequences relative to other CVB3 strains. Progeny CVB3/GA replicated similarly to the parental strain in three different cell cultures and was avirulent when inoculated into mice, causing neither pancreatitis nor myocarditis. Inoculation of mice with CVB3/GA protected mice completely against myocarditis and pancreatitis induced by cardiovirulent CVB3 challenge. The secondary structure predicted for the CVB3/GA domain II, a region within the 5' non-translated region that is implicated as a key site affecting the expression of a cardiovirulent phenotype, differs from those predicted for cardiovirulent and pancreovirulent CVB3 strains. This is the first report characterizing a cloned CVB3 genome from an avirulent strain.

The genome of a very virulent Marek's disease virus.

Here we present the first complete genomic sequence, with analysis, of a very virulent strain of Marek's disease virus serotype 1 (MDV1), Md5. The genome is 177,874 bp and is predicted to encode 103 proteins. MDV1 is colinear with the prototypic alphaherpesvirus herpes simplex virus type 1 (HSV-1) within the unique long (UL) region, and it is most similar at the amino acid level to MDV2, herpesvirus of turkeys (HVT), and nonavian herpesviruses equine herpesviruses 1 and 4. MDV1 encodes 55 HSV-1 UL homologues together with 6 additional UL proteins that are absent in nonavian herpesviruses. The unique short (US) region is colinear with and has greater than 99% nucleotide identity to that of MDV1 strain GA; however, an extra nucleotide sequence at the Md5 US/short terminal repeat boundary results in a shorter US region and the presence of a second gene (encoding MDV097) similar to the SORF2 gene. MD5, like HVT, encodes an ICP4 homologue that contains a 900-amino-acid amino-terminal extension not found in other herpesviruses. Putative virulence and host range gene products include the oncoprotein MEQ, oncogenicity-associated phosphoproteins pp38 and pp24, a lipase homologue, a CxC chemokine, and unique proteins of unknown function MDV087 and MDV097 (SORF2 homologues) and MDV093 (SORF4). Consistent with its virulent phenotype, Md5 contains only two copies of the 132-bp repeat which has previously been associated with viral attenuation and loss of oncogenicity.

Sequence and initial characterization of the U(L)10 (glycoprotein M) and U(L)11 homologous genes of serotype 1 Marek's Disease Virus.

The nucleotide sequence of the U(L)10 (glycoprotein M) and the U(L)11 homologs of Marek's Disease Virus 1 strain GA was determined. The U(L)10 open reading frame encodes a type III membrane protein of 424 amino acids that contains eight hydrophobic domains and two consensus N-linked glycosylation sites. The U(L)11 homologous gene encodes an 84 amino acid polypeptide, and contains a highly conserved myristylation site at its aminoterminus. By analysis of infected-cell RNA with strand-specific RNA probes, transcription of both U(L)10 and U(L)11 in infected cells was demonstrated. Coupled in vitro transcription-translation confirmed that the U(L)10 product is a 47 kD N-glycosylated viral protein that aggregated upon boiling, whereas the U(L)11 protein exhibited a size of 12 kD after in vitro translation.

Selenomonas lipolytica sp. nov., an obligately anaerobic bacterium possessing lipolytic activity.

A novel, oligately anaerobic bacterium capable of hydrolysing lipids was isolated from a tropical anaerobic lagoon receiving waste water from an edible oil mill. The isolate had many characteristics similar to those of members of the genus Selenomonas. The isolate showed lipolytic activity on tributyrin, triolein and groundnut oil in qualitative plate clearance assays, which has not been reported for the type strain of the genus Selenomonas. It did not require n-valerate supplementation for growth on glucose. Acetate and propionate were the only volatile fatty acids produced from glucose fermentation with propionate as the major end product. The isolate could grow optimally at pH 6.8 and at a temperature of 40 degrees C. It could tolerate NaCl concentrations of up to 40 g l-1. The G&C content of the DNA was 40 mol% as determined by thermal denaturation analysis. Comparison of partial 165 rRNA gene sequences revealed that the isolate was most closely related to genus Selenomonas with 91% sequence similarity (250 bp compared) to Selenomonas ruminantium strain GA 192. On the basis of the results obtained in the present investigation, it is suggested that a new species of Selenomonas should be created for this novel isolate and the name Selenomonas lipolytica is proposed for this new species. The type strain is strain CF1BT (= MCMB 505T).

Isolation of Actinomyces viscosus strain GA: characteristics of its cellular biological retardant(s).

Actinomyces viscosus strain GA produces an exocellular biological retardant(s) that prevents certain vegetable plants from becoming overgrown. The biological retardant(s) was assayed using the etiolated wheat coleoptile assay, and fractionation of culture supernatant fluid resulted in a partial purification of the retardant(s). The biological retardant(s) was most active around pH 7 in the bioassay and when applied to sterile soil mixture. The biological retardant(s) was tentatively identified as a derivative of a rare hexose carbohydrate (but not an amino sugar) but an exact structure was not determined. In a sterilized synthetic soil system, the biological retardant(s) has an effect on tomato cultivars similar to that observed by the synthetic plant growth regulators Alar (succinic acid 2,2,-dimethylhydrazide) and Bonzi (paclobutrazol).

Characterization and expression of the Marek's disease virus serotype 2 glycoprotein E in recombinant baculovirus-infected cells: initial analysis of its DNA sequence and antigenic properties

In Marek's disease virus (MDV) serotype 2 (MDV2) genome, a gene equivalent to the glycoprotein E (gE) of other alphaherpesviruses was identified and sequenced. The primary translation product comprises 488 amino acids with a M r of 54.3 kDa. The predicted amino acid sequence possesses several characteristics typical of membrane glycoproteins, including a N-terminal hydrophobic signal sequence, C-terminal transmembrane and cytoplasmic domains, and extra-cellular region containing four potential N-linked glycosylation sites. Compared with other MDV serotypes, MDV2 gE showed 47.3% identity with MDV1 gE, and 38.9% identity with HVT gE at the amino acid level. In transcriptional analyses, a 2.0 kb mRNA which starts between 65 and 86 bps upstream of the potential translational initiation codon of gE was identified as the gE-specific transcript. By a recombinant baculovirus, this potential gE coding region was expressed as several specific products from 66 to 72 kDa. These products were susceptible to tunicamycin treatment, indicating that they were glycoprotein in nature. Further, the expressed gE reacted with all chicken-antisera raised to each of the three serotypes of MDV (strains GA, SB-1, and FC126), suggesting that gE is expressed by all three serotypes of MDV in infected cells and conserves common antigenic epitope(s) beyond those that are serotype specific.

2D–3D transition in highly strained [formula omitted] heterostructures by transmission electron microscopy

The 2D–3D transition occurring in highly strained GaAs Ga 1 - x In x As heterostructures growns by MBE was analyzed by specific TEM techniques and correlated to photoluminescence data. HREM evidences that the transition always happens in a standard growth for x ∼ 0.35. This leads to a purely elastic relaxation which is characterized by a sinusoidal modulation. When the temperature is decreased, it was confirmed by “average intensity profiles” recorded from digitized HREM images that the transition is pushed away towards greater thicknesses. Furthermore, when adding Te as a surfactant, 3 1 1 (g, ng) weak-beam images demonstrate that the 2D growth mode is maintained up to the plastic relaxation. Finally, the change in roughness and abruptness of the interfaces is discussed as a function of growth parameters.

Activation and degradation of benzoate, 3-phenylpropionate and crotonate by Syntrophus buswellii strain GA. Evidence for electron-transport phosphorylation during crotonate respiration

Activation and degradation of benzoate, 3-phenylpropionate and crotonate by Syntrophus buswellii strain GA. Evidence for electron-transport phosphorylation during crotonate respiration

Activation and degradation of benzoate, 3-phenylpropionate and crotonate by Syntrophus buswellii strain GA. Evidence for electron-transport phosphorylation during crotonate respiration.

A strictly anaerobic, benzoate-degrading bacterium, Syntrophus buswellii strain GA, was able to degrade benzoate or 3-phenylpropionate to acetate, CO2 and H2, if the hydrogen partial pressure was sufficiently low. The hydrogen was removed in syntrophic coculture by Methanospirillum hungatei or by Desulfovibrio sp. through interspecies hydrogen transfer or in pure culture by the use of crotonate as reducible cosubstrate. Alternatively, S. buswellii strain GA could grow in pure culture with crotonate. Activities of seven catabolic enzymes were measured in crude cell extracts of S. buswellii strain GA grown with various substrates and of crotonate-grown S. buswellii strain DSM 2612A. Benzoate, 3-phenylpropionate and crotonate were activated by CoA ligases. Glutaryl-CoA dehydrogenase was found to be involved in the degradation of aromatic compounds and enzymes catalysing beta-oxidation were involved in the reaction sequence from crotonyl-CoA to acetate. A c-type cytochrome was present in the cytoplasm, whereas b-type cytochromes were associated with the membranes of both S. buswellii strains grown on crotonate. These indicated the presence of an electron-transport system. A high growth yield of crotonate-grown S. buswellii strain GA might be explained by electron-transport phosphorylation in addition to substrate-level phosphorylation.

Marek's Disease Virus Serotype 2 Glycoprotein I Gene: Nucleotide Sequence and Expression by a Recombinant Baculovirus.

In the Marek's disease virus (MDV) serotype 2 (MDV2) genome, a gene equivalent to the glycoprotein I (gI) of other alphaherpesviruses was identified and sequenced. The primary translation product comprises 355 amino acids with a M(r) of 38.4 kDa. The predicted amino acid sequence possesses several characteristics typical of membrane glycoproteins, including a N-terminal hydrophobic signal sequence, C-terminal transmembrane and cytoplasmic domains, and extra-cellular region containing three potential N-linked glycosylation sites. Compared to other MDV serotypes, MDV2 gI showed 49% identity with MDV1 gI, and 36% identity with HVT gI at the amino acid level. In transcriptional analyses, a 3.5 kb mRNA which starts between 56 and 147 bps upstream of the potential translational initiation codon of gI was identified as the gI-specific transcript. By a recombinant baculovirus, this potential gI encoding region was expressed as two specific products 45 and 43 kDa. Both products were susceptible to tunicamycin treatment, indicating that they were glycoprotein. Further, the expressed gI reacted with all chicken-antisera raised to each of the three serotypes of MDV (strains GA, SB-1, and FC126), suggesting that gI is expressed by all three serotypes of MDV in infected cells and conserves common antigenic epitope(s) beyond serotypes.