Xanthomonas oryzae pv. oryzae XopQ protein suppresses rice immune responses through interaction with two 14-3-3 proteins but its phospho-null mutant induces rice immune responses and interacts with another 14-3-3 protein.

Many bacterial phytopathogens employ effectors secreted through the type-III secretion system to suppress plant innate immune responses. The Xanthomonas type-III secreted non-TAL effector protein Xanthomonas outer protein Q (XopQ) exhibits homology to nucleoside hydrolases. Previous work indicated that mutations which affect the biochemical activity of XopQ fail to affect its ability to suppress rice innate immune responses, suggesting that the effector might be acting through some other pathway or mechanism. In this study, we show that XopQ interacts in yeast and in planta with two rice 14-3-3 proteins, Gf14f and Gf14g. A serine to alanine mutation (S65A) of a 14-3-3 interaction motif in XopQ abolishes the ability of XopQ to interact with the two 14-3-3 proteins and to suppress innate immunity. Surprisingly, the S65A mutant gains the ability to interact with a third 14-3-3 protein that is a negative regulator of innate immunity. The XopQS65A mutant is an inducer of rice immune responses and this property is dominant over the wild-type function of XopQ. Taken together, these results suggest that XopQ targets the rice 14-3-3 mediated immune response pathway and that its differential phosphorylation might enable interaction with alternative 14-3-3 proteins.

Design of Experiments (DoE) Approach to Optimize the Sustained Release Microparticles of Gefitinib.

BACKGROUND: Gefitinib (GEF), the kinase inhibitor, is presently available as tablets to be taken orally in high doses of 250-500 mg per day due to its poor solubility. The solubility issues affect not only its onset of action but also the bioavailability. These drawbacks foresight the need to have an alternate dosage form, preferably a sustained release formulation. METHODS: In the present study, microparticles were prepared by emulsion solvent evaporation using PLGA 50:50 (GEF-PLGA MP). A 32 factorial design was used to optimize the critical quality parameters to the set mean particle size in the range of 7.4±2.5 µm and entrapment efficiency of 80%. SEM microscopy of the prepared microparticles confirmed to have a spherical smooth shape. The GEFPLGA- MPs sustained the release of GEF for 72 hours. The first-order kinetics ruled the mechanism of drug release and was predicted to follow Fickian diffusion. RESULT: Anticancer efficacy was judged by the cytotoxicity studies using the L132 lung cancer cells. MTT assay showed 3-fold enhanced cytotoxicity of GEF loaded microparticles against L132 cells as compared to plain GEF. CONCLUSION: It was concluded that gefitinib can be efficiently loaded into the biodegradable polymer PLGA to provide sustained release of the drug.

Diagnostic adequacy and safety of endoscopic ultrasound-guided fine-needle aspiration in patients with lymphadenopathy in a large cohort.

BACKGROUND AND STUDY AIMS: The role of endoscopic-ultrasound (EUS) guided fine-needle aspiration (FNA) in patients with lymphadenopathy in terms of diagnostic adequacy and safety in large population is not well defined. The aim of this study was to evaluate diagnostic adequacy and safety of EUS-FNA in patients with lymphadenopathy. PATIENTS AND METHODS: Retrospective study from October 2010 to September 2015 at tertiary care center in Delhi-NCR. We analyzed data from 1005 EUS- FNAs of lymph nodes. RESULTS: The study cohort comprised 1005 lymph nodes in 865 patients; 68 % were males, mean age was 50 ±â€Š14 years. Indications of FNA were to look for etiology of pyrexia of unknown origin or staging of malignancy mainly. FNA was taken from mediastinal nodes (n = 528, 52.5 %) and intra-abdominal nodes (n = 477, 47.5 %). Median size of nodes at long axis and short axis was 17 (12 - 25.7) and 10 (8 - 15) mm respectively. Adequate material by FNA was obtained in 92.8 % cases. The cytopathologic diagnosis were malignancy in 153 (15.2 %), granulomatous change in 452 (42 %), and reactive lymphadenopathy in 328 (35.6 %). There was statistically significant difference seen between groups with pathological and reactive lymph nodes regarding size at long and short axis, hypoechoic nature, well defined borders and presence of necrosis and calcification. Procedure-related adverse effects were encountered in 6 patients (0.8 %). Four patients had mild mucosal bleeding in chronic liver disease patients and two had mild hepatic encephalopathy related to sedation. CONCLUSION: EUS-FNA of lymph nodes has good diagnostic adequacy and safety.

Occurrence of a Clover Proliferation (16SrVI) Group Phytoplasma Associated with Little Leaf Disease of Portulaca grandiflora in India.

Portulaca grandiflora (family Portulacaceae), commonly known as moss rose purslane, is a popular ornamental plant widely grown in temperate climates because it blooms all summer. Portulaca is also used for medicinal purposes since it is rich in vitamins A, B1, and C and has antimicrobial and cytotoxic activity. Since March 2005, 30 to 50% of P. grandiflora plants in the ornamental gardens as well as in pots at the Central Institute of Medicinal and Aromatic Plants, Lucknow, India have displayed symptoms resembling phytoplasma infection. Disease symptoms start as a typical bud proliferation, downward curling, and diminishing size of leaves, followed by overall stunted growth and yellowing of the whole plant from April to June. Some plants also formed rosettes and a proliferation of axillary shoots resulting in a witches'-broom appearance. Typical pleomorphic bodies, mostly spherical to oval, ranging from 340 to 1,100 nm were observed only in sieve elements of infected plants by transmission electron microscopy (TEM). On the basis of symptoms, TEM observations, PCR, and response to antibiotic treatment, the causal organism was identified as phytoplasma (1). Total genomic DNA from healthy and infected plants was extracted with the CTAB buffer method (2). Of 27 suspected samples screened by PCR, 23 were phytoplasma positive. Presence of phytoplasmas in plants was demonstrated by a nested PCR assay employing primer pair P1/P6 followed by R16F2n/R16R2 that generated rDNA products of 1.5 and 1.2 kb, respectively, only from symptomatic plants. No differences among phytoplasmas in Portulaca plants were detected by restriction fragment length polymorphism (RFLP) analysis of nested rDNA (1.2 kb) products using endonucleases BamHI, RsaI, AluI, HpaII, and EcoRI. Comparative analysis of RFLP patterns with those derived from reference phytoplasmas tentatively identified the Portulaca little leaf (PLL) phytoplasma as a member of 16S rDNA RFLP group 16SrVI (3). A nested PCR product (1.25 kb) was cloned with a TOPO TA cloning kit (Invitrogen, Carlsbad, CA) and sequenced. The sequence was deposited in the GenBank database (Accession No. EF651786). Sequence analysis revealed the PLL phytoplasma to be most similar (98%) to Indian brinjal little leaf (Accession No. EF186820) and 'Candidatus Phytoplasma trifolii' (Accession No. AY390261), two 16SrVI group phytoplasmas previously reported from India and Canada, respectively. The status of PLL (EF651786) was also verified by in silico RFLP analysis (4) of the F2n/R2 sequence of six closely related strains (Accession Nos. AF228052, AY390261, AY270156, AY409070, AY409069, and EF186820) of the 16SrVI group using 17 restriction enzymes (AluI, BamHI, BfaI, BsfUI, DraI, EcoRI, HaeIII, HhaI, HinfI, HpaI, HpaII, KpnI, MseI, Sau3AI, RsaI, SspI, and TaqI). In silico restriction digestion and virtual gel plotting showed similar patterns for all enzymes. To our knowledge, this is the first report of a 16SrVI group phytoplasma infecting Portulaca plants in India. References: (1) P. V. Ajayakumar et al. Aust. Plant Dis. Notes 2:67, 2007. (2) S. P. S. Khanuja et al. Plant Mol. Biol. Rep. 17:74, 1999. (3) I.-M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998. (4) W. Wei et al. Int. J. Syst. Evol. Mic. 57:1855, 2007.