Unfamiliar necrotic symptoms on or within potato tubers of cultivars Nishiyutaka and Dejima were observed in Nagasaki prefecture, Japan, in 1992. Symptoms were typically on the surface of the tuber, which either protruded at first and then became sunken, or showed necrotic spots, with necrosis within the tubers. Symptoms sometimes appeared at harvesting but more often appeared after storage for several months. Investigations revealed that the causal agents of the disease were isolates of Potato virus Y necrotic strain (PVY^NTN) and the disease was potato tuber necrotic ringspot disease (PTNRD), previously reported in Europe and Lebanon. Five potato cultivars were inoculated with an isolate from necrotic tubers. The highest percentage of progeny tubers showing PTNRD was found in cv. Nishiyutaka (23.3%). In contrast, cvs. Shima-bara, Mayqueen, and Danshaku showed a low percentage of PTNRD. Additional potato tubers with PTNRD were also observed after storage of the tubers. To investigate the relatedness between isolates of PVY^NTN and of necrotic strain PVY^N, previously isolated in Japan, Nishiyu-taka was inoculated with an isolate of PVY^N, which also induced PTNRD. Nucleotide sequences of coat protein (CP) genes of six PVY^NTN isolates were determined. The CPs were 267 amino acids in length, with a substitution of one or no amino acid among each of the six isolates. The phylogenetic relationship based on nucleotide sequences of CP genes showed that these six PVY^NTN isolates clustered together with PVY^N isolates. This is the first report of PTNRD caused by PVY^NTN isolates in Asia.

The nephron site responsible for the different patterns of sodium excretion in response to extracellular volume expansion observed in dogs with bilateral acute remnant kidneys, acute NTN, and normal kidneys was studied with clearance and micropuncture techniques. Mean kidney GFR's in the remnant and NTN kidneys were similar at 14 and 15 ml/min, respectively, compared to 28 ml/min for the normal kdineys. However, SNGFR's were normal in the remnant kidneys but markedly reduced in dogs with NTN. Mean absolute sodium excretion was similar for the remnant and normal kidneys both during the control phase and after volume expansion. Because of the reduced GFR, FENa of the remnant kidneys was significantly higher in each situation. In contrast, mean absolute sodium excretion was markedly less in dogs with NTN than in normal dogs both before and after volume expansion. Although FENa during the control phase was similar to that in normal dogs, it increased significantly less with volume expansion. Despite these differences in urinary sodium excretion, the percent sodium reabsorption at the end of the proximal convoluted tubule was similar in all three groups. In addition, volume expansion depressed proximal SFENa to the same degree in each group. Therefore the different patterns of urinary FENa were the result of differences in fractional sodium reabsorption by the nephron segments distal to the proximal convoluted tubule. Decreased distal delivery of sodium secondary to the reduced SNGFR also contributed to the decreased sodium excretion in acute NTN.

BACKGROUND

Quinoline-3-carboximide compounds, such as paquinimod, which targets the protein S100A9, have demonstrated efficacy in treating autoimmune diseases. S100A9, in association with S100A8, forms the heterodimer S100A8/S100A9, known as calprotectin; that has been shown to be upregulated in numerous inflammatory disorders. We had previously demonstrated protection from glomerular disease in S100A9-deficient mice. The aim of this study was to assess the efficacy of paquinimod in the prevention and treatment of experimental glomerulonephritis.

METHODS

Nephrotoxic nephritis (NTN) was induced in C57BL/6 mice according to our standard protocol. Mice were treated with different doses of paquinimod either at disease induction (prevention group) or two days following induction (therapeutic group) and sacrificed 8 days following induction. Disease was assessed histologically (number of glomerular crescents, degree of glomerular thrombosis, number of infiltrating leucocytes and calprotectin expression) and biochemically (serum creatinine and urea levels, and urinary levels of protein).

RESULTS

Neither treatment with low (0.5mg/kg) or high (25mg/kg) doses of paquinimod, given preventatively or therapeutically, led to disease attenuation, as assessed by biochemical or histological parameters. Additionally, we found trends for an increase in renal glomerular calprotectin expression in the high dose groups, suggesting a possible feedback regulation of calprotectin expression.

CONCLUSIONS

Our results show that paquinimod does not successfully prevent or treat mice with NTN. Other models of immune-mediated glomerulonephritis need to be tested to investigate the therapeutic potential of this compound in renal disease.

The importance of potato has increased dramatically in Indonesia over the last three decades. During this period, 'Granola', a potato cultivar originally from Germany, has become the most common cultivar for fresh consumption in Indonesia. In August 2014, a survey was conducted in Sulawesi, where potato fields cultivated with Granola and its selection, 'Super John', were sampled for Potato virus Y (PVY) presence. PVY was found in Sulawesi for the first time. Samples determined to be positive for PVY were subsequently typed to strain using reverse-transcription polymerase chain reaction assays. All PVY isolates sampled were identified as PVY^NTN recombinants, with three recombination junctions in P3, VPg, and CP regions of the genome. Three local PVY isolates were subjected to whole-genome sequencing and subsequent sequence analysis. The whole genomes of the Indonesian PVY^NTN isolates I-6, I-16, and I-17 were found to be closely related to the European PVY^NTN-A. This recombinant type was shown previously to cause potato tuber necrotic ringspot disease (PTNRD) in susceptible potato cultivars. The dependence of potato farmers on mostly a single cultivar, Granola, may have given a competitive advantage to PVY^NTN over other PVY strains, resulting in the predominance of the PVY^NTN recombinant. The dominance of PVY^NTN in Sulawesi, and possibly in Indonesia as a whole, represents a potential risk to any newly introduced potato cultivar to the country, especially cultivars susceptible to PTNRD.

A presynthesized, square planar copper imidazole complex, [Cu(imidazole)4](NO3)2, was utilized as a precursor in the synthesis of a new series of zeolitic imidazolate frameworks, termed ZIF-202, -203, and -204. The structures of all three members were solved by single-crystal X-ray diffraction analysis, which revealed ZIF-203 and -204 having successfully integrated square planar units within the backbones of their respective frameworks. As a result of this unit, the structures of both ZIF-203 and -204 were found to adopt unprecedented three-dimensional nets, namely, ntn and thl, respectively. One member of this series, ZIF-204, was demonstrated to be highly porous, exhibit exceptional stability in water, and selectively capture CO2 over CH4 under both dry and wet conditions without any loss in performance over three cycles. Remarkably, the regeneration of ZIF-204 was performed under the mild conditions of flowing a pure N2 gas through the material at ambient temperature.

The roles of polyhydroxy-butyrates/alkanoates (PHB/PHA) in biology, for the preparation of chiral building blocks, and as a source of inspiration for the discovery of β- and γ-peptides are discussed. The syntheses and structures of β-peptides are outlined. The prerequisites for mimicking peptide/protein interactions with β-peptides and two examples are presented. Single terminal β-amino-acid residues can lead to stabilization of peptides (cf. NTS(8-13)) in plasma. Cell-penetrating α-L-, α-D-, mixed α-L/D- and β-oligoarginines (OAs) and -oligoprolines, as well as the mechanism(s) of internalization are compared. Recent studies show that infected erythrocytes, parasitic organisms and mycobacteria are entered by OA-derivatives, which have been employed as transporters of the antibiotic fosmidomycin. While β-peptides are generally enzymatically stable (for days in mammals), a microorganism (S. xenopeptidilytica) with an Ntn enzyme (3-2W4 BapA) was discovered that cleaves only β-peptides, and that was applied in preparations of (enantiopure) β-amino acids and β-peptides.

An isolate of potato virus Y (PVY), PVY-M3, was subjected to biological characterization on potato indicators and to whole-genome sequencing. PVY-M3 induced a local and systemic hypersensitive resistance (HR) response in potato cultivar Maris Bard expressing the Nz gene while inducing no HR in potato cultivars Desiree and King Edward, carrying Ny and Nc genes, respectively. These HR responses, combined with a lack of vein necrosis in tobacco, clearly defined PVY-M3 as an isolate of the PVY(Z) strain. Recombination analysis demonstrated that PVY-M3 had a typical European PVY(NTN) genome with three recombinant junctions, and PVY(N) and PVY(O) were identified as parents.

Potato tuber necrotic ringspot disease (PTNRD), caused by potato Y potyvirus isolate NTN (PVYNTN) of the N virus strain group, was first described in Hungary in 1980. It has spread throughout Europe, with the most recent reports from Portugal and Italy in 1997 to 1998 (1). Superficial necrotic ringspot areas on tubers, typical of PTNRD, were first observed in commercial potato fields of the Nevrokopi region in northern Greece in 1994. Affected cultivars included Timate and, to a lesser extent Spunta, the original seed of which came from the Netherlands. PVY was identified from all tubers tested by indexing on Nicotiana tabacum and subsequent testing of the plants by double antibody sandwich-enzyme-linked immunosorbent assay (DAS-ELISA). The potato seed lots were rejected by the local authorities. PTNRD reappeared in the same area in 1998 in a more aggressive manner. Cultivar Hermes, imported from Scotland, was most affected, with very severe symptoms in 80% of the tubers. Symptoms appeared in early September, 40 days after defoliation of the plants. Other cultivars were affected at lower rates; cv. Spunta showed typical symptoms and cvs. Fabola, Santana, and Irvila exhibited atypical cracks and blisters. In all cases, PVY was isolated in N. tabacum and its presence confirmed by DAS-ELISA. Immunocapture-reverse transcription-polymerase chain reaction with specific PVYNTN primers (2) detected the characteristic 835-bp product in all cultivars tested. The incidence of PTNRD seems to be expanding in northern Greece, where it has become a threat to potato production. PTNRD symptoms were also observed in southern Greece (Ahaia) in experimental "crossing" fields of seed stocks. In this case, the disease seems to have spread especially in cv. Marfona. References: (1) L. Tomassoli et al. Plant Dis. 82:350, 1998. (2) H. L. Wiedemann and E. Maiss. Z. Pflanzenkrankh. Pflanzenschutz 103:337, 1996.

Most strains of Potato virus Y (PVY) can infect tobacco plants (Nicotiana tabacum) and cause vein clearing followed by leaf mottling, except the PVY^N strain, which induces severe vein necrosis. Some isolates within the PVY^N strain also cause potato necrotic tuber ringspot disease, but these have not been reported from Canadian tobacco fields. PVY^NTN isolates include European (EU) and North American (NA) types that are serologically identical to PVY^N, but can be distinguished by nucleic acid-based assays and potato bioassay (1,2). Some PVY isolates, PVY^N-Wi or PVY^N:O, resulting from a recombination between RNA molecules of PVY^N and the common strain, PVY^O, are identified as PVY^O in serological assays, but induce necrosis in tobacco (2). In August of 2007, two samples of tobacco (N. tabacum, unknown cultivar) leaves showing necrotic symptoms resembling those induced by PVY^N, PVY^NTN, or PVY^N-Wi were collected from a tobacco field in southern Ontario, Canada and submitted to the Charlottetown Laboratory, Canadian Food Inspection Agency, Charlottetown, PE. Virus in both samples (PVY-204 and PVY-205) reacted with PVY^N-specific antibodies 1F5 and 4E7 (3) and induced vein necrosis in tobacco (N. tabacum cv. Samsun). A multiplex reverse transcription (RT)-PCR assay (1) for the simultaneous detection and differentiation of various PVY strains amplified two fragments (181 and 452 bp) associated with EU-PVY^NTN isolates. Restriction fragment length polymorphism (RFLP) analysis targeting the P1 and NIb gene (3) also indicated that PVY-204 and PVY-205 were EU-PVY^NTN isolates. Known isolates of PVY^O, PVY^N, and NA-PVY^NTN were used in all evaluations as references (3). Furthermore, the nucleotide sequences of the P1 and NIb genes of PVY-204 and PVY-205 determined by automated cycle sequencing (3) and subjected to phylogenetic analysis indicated that the nucleotide and deduced amino acid sequences of both isolates were 96 and 95% identical, respectively, to NA-PVY^NTN isolates reported from Canada, but 99% identical (both nucleotide and amino acid) to EU-PVY^NTN isolates from Europe and Mexico (3). Potato (cv. Yukon Gold) plants mechanically inoculated with leaf sap from tobacco (N. tabacum cv. Samsun) infected with PVY-204 and PVY-205 developed various leaf symptoms including severe local and systemic necrotic lesions, leaf wilting, and leaf death in 3 to 5 weeks postinoculation under greenhouse conditions. The infected plants recovered in 5 to 6 weeks. Potato (cv. Yukon Gold) plants inoculated with leaf sap from tobacco (N. tabacum cv. Samsun) infected with a PVY^NTN isolate (HX8) (3) and healthy tobacco leaf sap were used as positive and negative controls. The number and yield of the tubers harvested from infected plants were significantly reduced (50%), and PVY-204 and PVY-205 induced typical potato tuber necrotic ringspot disease in 52.6% of the progeny tubers with an average disease index of 0.364 (C. Kerlan and K. Charlet-Ramage, EAPR Virology Meeting Proceedings, 1998). PVY^NTN was detected by RT-PCR and RFLP in all necrotic tubers and 66.7% of the asymptomatic tubers. Some tubers (15.8%) harvested from the infected plants were negative in RT-PCR targeting either P1 protein gene or NIb gene and showed neither external nor internal necrotic symptom. To my knowledge, this is the first evidence of the occurrence of PVY^NTN isolates in field-grown tobacco plants in Canada. References: (1) J. H. Lorenzen et al. Plant Dis. 90:935, 2006. (2) R. Singh et al. Arch Virol. 153:1, 2008. (3) H. Xu et al. Can. J. Plant Pathol. 27:125, 2005.

Plant viral infections lead to accumulation of virus-derived small interfering RNAs (vsiRNAs) as a result of host defense mechanisms. High-throughput sequencing technology enables vsiRNA profiling analyses from virus infected plants, which provide important insights into virus-host interactions. Potato virus Y (PVY) is a detrimental plant pathogen that can infect a variety of solanaceous crops, e.g., potato, tobacco, tomato, and pepper. We analyzed and characterized vsiRNAs derived from Nicotiana tabacum cv. Samsun infected with two recombinant PVY strains, N-Wi and NTN. We observed that the average percentage of vsiRNAs derived from plants infected with N-Wi was higher than from plants infected with NTN, indicating that N-Wi invokes a stronger host response than NTN in tobacco. The size distribution pattern and polarity of vsiRNAs were similar between both virus strains with the 21 and 22 nucleotide (nt) vsiRNA classes as most predominant and the sense/antisense vsiRNAs ratio nearly equal in the 20-24 nt class. However, the percentage of sense vsiRNAs was significantly higher in the 25-26 nt long vsiRNAs. Distinct vsiRNA hotspots, identifying highly abundant reads of different unique vsiRNA sequences, were observed in both viral genomes. Previous studies found an A or U bias at the 5' terminal nucleotide position of 21 nt vsiRNAs; in contrast, our analysis revealed a C and U nucleotide bias. This study provides insights that will help further elucidate differential processing of vsiRNAs in plant antiviral defense.

Keywords: Potato virus Y; Small RNA profiling; Tobacco; vsiRNA.

Surveys of commercial and seed potato fields for virus diseases (1998 to 2002) in Manitoba established that Potato virus Y (PVY) is of concern in seed potato production. To determine the prevalence of PVY strains, PVY-infected tubers identified by reverse transcription-polymerase chain reaction (RT-PCR) from surveys (2000 to 2001) were grown for symptom expression and strain characterization by strain-specific RT-PCR, bioassays, and serological assays. Of the samples collected (2000 to 2001) and tested by RT-PCR, 4.0% contained PVY. Further analysis of the PVY-positive samples by a duplex RT-PCR facilitating the simultaneous detection of common (PVY^O) and tobacco veinal necrosis strains (PVY^N/NTN) indicated that 37.5% contained PVY^O and 63.5% contained PVY^N-type isolates. Analysis of the PVY^N-type samples using three monoclonal antibodies (MAbs) showed that all reacted with only the PVY^O MAbs and not with the PVY^N-specific MAb. Partial nucleotide sequences of both ends of PVY-RNA showed that the PVY^N-type isolates resembled those reported in 1996 from Manitoba. These isolates are designated as PVY^N:O. In view of the increased incidence of PVY^N:O in one production area, seed tubers imported from other provinces of Canada and the neighboring United States were analyzed for PVY^N:O. The PVY^N:O was detected in imported seeds from Minnesota, Montana, and North Dakota.

The present study demonstrates how different potato virus Y (PVY) strains affect the miRNA balance in tobacco cv. Samsun. The two prevalent strains PVYNTN and PVYN-Wi caused severe and mild veinal necrosis (VN) respectively, and the unique PVYZ-NTN strain induced milder vein clearing (VCl) in the upper non-inoculated leaves. A single amino acid polymorphisms (SAPs) I252V and a Q412 to R412 substitution in the HC-Pro cistron of the PVYZ-NTN strain might relate to the loss of VN in tobacco. The abundance of 18 out of the 26 tested miRNAs was increased upon infection by the severe strains PVYNTN and PVYN-Wi. Expression of a group of defense related transcripts were increased accordingly. Two miRNAs, nta-miR6020a-5p and nta-miR6164a/b, which target the TIR-NBS-LRR type resistant TMV N genes involving in signal transduction, might correlate with the PVYNTN and PVYN-Wi induced VN. The down-regulated mRNAs, e.g., RAP2-7 and TOE3, PXC3, LRR-RLK, ATHB-14 and TCP4 targeted by nta-miR172, nta-miR390, nta-miR482, nta-miR166 and nta-miR319/159 respectively, were related to regulation of transcription, protein phosphorylation and cell differentiation. The observed strain-specific alteration of miRNAs and their targets are host dependent and corresponds to the symptom severity and the viral HC-Pro RNA levels.

Potato virus Y (PVY) has been reported in potato crops in Mexico (3), with tobacco necrotic variants found in the central State of Mexico (4). Nevertheless, many individual states are currently declared PVY free and distribution of individual strains of PVY in potato in different states of Mexico and in different solanaceous crops had not yet been studied. A limited field PVY survey was conducted on potato in the State of Chihuahua in August 2009. More than 900 random potato leaf samples were collected from cvs. Snowden, Atlantic, FL1867, Felsina, Fianna, Gigant, and Alpha. Seven were found to be PVY-positive and had been collected from cvs. Fianna, Snowden, and FL1867. The PVY status of the collected samples was initially determined with the PVY-specific Immunostrips (Bioreba, Reinach, Switzerland) and by double-antibody sandwich-ELISA using the polyclonal PVY detection kit (Agdia, Elkhart, IN). To determine the strain specificity of these PVY isolates following ELISA tests, the infected original samples were inoculated onto tobacco plants at the four-leaf stage and symptom appearance and development were observed for 8 weeks side-by-side with control isolates PB-Oz (PVY^O), N4 (PVY^NTN), and Mont (PVY^N) (1), followed by the standard PVY strain typing by reverse transcription (RT)-PCR (2). Only one of the PVY-positive samples, originally from symptomless potato cv. Fianna, induced systemic PVY infection in tobacco by producing stunting, mosaic, and vein clearing. No systemic vein necrosis, characteristic of isolates Mont and N4, was observed in Nicotiana tabacum cvs. Burley, Xanthi, or Samsun after inoculation with this isolate during all 8 weeks of observation. This isolate, PVY-M3, was typed as a PVY recombinant by RT-PCR, with two recombinant junctions characteristic of European PVY^NTN strains (2). It was further analyzed by triple-antibody sandwich-ELISA using four PVY^O and PVY^N strain-specific monoclonal antibodies. Monoclonals 1F5 (Agdia) and SASA-N (Scottish Agriculture Science Agency [SASA], Edinburgh) reacted to this isolate and identified PVY-M3 serologically as PVY^N serotype, characteristic of other PVY^NTN recombinants. Monoclonals MAb2 (Agdia) and SASA-O (SASA), specific to PVY^O and PVY^C strains, did not react to PVY-M3. Taken together, the combination of biological, serological, and molecular characteristics define this recombinant isolate from Mexico as belonging to the same PVY strain group represented by the isolate PVY-L26 (1). To our knowledge, this is the first report of such an unusual PVY^NTN recombinant strain from Mexico. Presence of this isolate, with no vein necrotic symptoms induced on tobacco and with PVY^NTN genome, will necessitate development of new detection methods for the seed potato industry in Mexico. References: (1) X. Hu et al. Virus Res. 143:68, 2009. (2) J. L. Lorenzen et al. Plant Dis. 90:935, 2006. (3) L. P. Moreno et al. Rev. Mex. Fitopatol. 22:187, 2004. (4) V. R. Ramirez-Rodriguez et al. Virol. J. 6:48, 2009.

Potato virus Y (PVY) causes substantial losses in potato production by decreasing yields and affecting the quality of potato tubers. Management of PVY in potato is dependent primarily on potato seed certification programs to prevent or limit initial levels of virus inoculum. Prior to 1990, the ordinary strain of PVY (PVY^O) was the predominant virus in North America. PVY^O induces clear foliar symptoms in many potato cultivars, allowing successful management in seed potato through a combination of visual inspections and limited laboratory testing. In recent years, necrotic strains of PVY (PVY^N, PVY^NTN, and PVY^N:O) have begun to spread in the United States, many of which induce mild symptoms in potato, making them more difficult to manage through visual inspections. In addition to reducing yield, necrotic isolates may also cause external and internal damage in tubers of susceptible cultivars, which is known as potato tuber necrotic ringspot disease (PTNRD). Tuber necrotic strains of PVY have been reported across the northern United States (1,2,4), although limited information is available on their incidence and spread in commercial potato production. During June and July of 2007, 38 random samples were collected from three different commercial fields displaying disease problems (cvs. Russet Ranger, Alturas, and Russet Burbank) in the vicinity of Idaho Falls, ID. Plants collected showed various degrees of mosaic and leaf yellowing. By using double-antibody sandwich (DAS)-ELISA and reverse transcription (RT)-PCR, 25 of these plants were identified as PVY positive. The mutiplex RT-PCR assay (3) confirmed that nine plants were infected with PVY^NTN and 11 with PVY^N:O. No RT-PCR products were amplified from five samples. During September and October of 2007, 25 tuber samples (cv. Russet Burbank) showing various degrees of unusual internal symptoms (e.g., brown spots) were collected near Idaho Falls, ID. Twenty-two tubers were found PVY positive by DAS-ELISA, and multiplex RT-PCR determined 13 of those were PVY^NTN, three were PVY^O, one was a PVY^NTN/N:O mixture, and one was a PVY^O/N:O mixture. No RT-PCR products were amplified from four samples. In October 2007, six tubers showing distinct external tuber damage characteristic of PTNRD (cv. Highland Russet) were collected near Twin Falls, ID. All six tubers were determined to be PVY positive by DAS-ELISA, and RT-PCR identified five as infected with PVY^NTN and one with PVY^N:O. All the mixtures were easily separated by inoculating tobacco plants followed by subsequent testing of individual plants. Asymptomatic tubers from the same lot not showing PTNRD damage were found PVY negative by DAS-ELISA and RT-PCR. All PVY^NTN isolates collected during 2007 were inoculated into tobacco plants (Nicotiana tabacum L. cv. Xanthi) and confirmed to induce systemic vein necrosis. Limited sequencing of four of the PVY^NTN isolates determined that they contained recombinant junctions 2 and 3, identifying them as being related to the European strain of PVY^NTN (3). The data suggest an increase in distribution and incidence of necrotic strains of PVY in commercial, potato-production areas in Idaho during an outbreak in 2007 and the potential for an increase in PTNRD. References: (1) P. M. Baldauf et al. Plant Dis. 90:559, 2006. (2) J. M. Crosslin et al. Plant Dis. 90:1102, 2006. (3) J. H. Lorenzen et al. Plant Dis. 90:935, 2006. (4) L. M. Piche et al. Phytopathology 94:1368, 2004.

The responses of 14 potato cultivars to five Potato virus Y (PVY) isolates belonging to four strains (ordinary [PVY^O], tobacco veinal necrosis [PVY^N], N:O group [PVY^N:O], and nonrecombinant potato tuber necrotic [PVY^NTN]) were studied in primary and secondary infections. For the primary infection experiments, foliage symptoms were monitored daily after mechanical inoculation with a PVY isolate until harvest; and, for the secondary infection experiments, foliage symptoms were monitored regularly from plant emergence until harvest. Tuber symptoms (namely, tuber necrotic ringspots) were checked at harvest and monthly postharvest for up to 4 months. In both infections, symptoms varied significantly depending on potato cultivar and virus strain or isolate. In primary infections, local lesions occurred on inoculated leaves of 'AC Chaleur', 'Eramosa', 'Goldrush', 'Jemseg', 'Katahdin', 'Ranger Russet', and 'Yukon Gold' after inoculation with PVY^O isolates, followed by systemic necrosis on latterly emerged uninoculated leaves. In contrast, plants of 'CalWhite', 'La Rouge', 'Red LaSoda', 'Russet Burbank', 'Russet Norkotah', and 'Superior' did not exhibit any visible symptoms on inoculated leaves but developed mild to severe mosaic on latterly emerged leaves after infection with PVY^O isolates. In all cultivars, near-symptomless to mild mosaic was induced by PVY^N and mild to severe mosaic by PVY^N:O. PVY^NTN induced mild to severe mosaic in plants of all cultivars except AC Chaleur, 'Cherokee', and Yukon Gold, which developed visible systemic necrosis. Necrotic ringspots were observed in tubers of PVY^NTN-infected plants of AC Chaleur, Cherokee, and Yukon Gold. The tuber symptoms were also incited by PVY^N-Jg on Cherokee. In secondary infections, the symptoms were generally more severe than primary infections even though the symptom types did not alter. As in the greenhouse, a clear symptom severity pattern (PVY^O-FL > PVY^O-RB > PVY^NTN-Sl > PVY^N:O-Mb58 > PVY^N-Jg) was observed in AC Chaleur, Cherokee, Eramosa, Goldrush, Jemseg, Katahdin, Ranger Russet, and Yukon Gold in the field.

Lupus nephritis (LN) is a serious end organ complication of systemic lupus erythematosus. Nephrotoxic serum nephritis (NTN) is an inducible model of LN, which utilizes passive transfer of pre-formed nephrotoxic antibodies to initiate disease. In previous studies, we demonstrated that the Bruton's tyrosine kinase inhibitor, BI-BTK-1, prevents the development of nephritis in NTN when treatment was started prior to nephrotoxic serum transfer, and reverses established proteinuria as well. We manipulated the initiation and duration of BI-BTK-1 therapy in NTN to study its delayed therapeutic effects when treatment is given later in the disease course, as well as to further understand what effect BI-BTK-1 is having to prevent initiation of nephritis with early treatment. Early treatment and remission induction each correlated with decreased inflammatory macrophages, CD4+ and CD8+ T cells, and decreased B220+ B cells. Additionally, an increased proportion of resident macrophages within the CD45+ population favored a delay of disease onset and remission induction. We also studied the cellular processes involved in reactivation of nephritis by withdrawing BI-BTK-1 treatment at different time points. Treatment cessation led to either early or later onset of renal flares inversely dependent on the initial duration of BTK inhibition, as assessed by increased proteinuria and BUN levels and worse renal pathology. These flares were associated with an increase in kidney CD45+ infiltrates, including myeloid cell populations. IL-6, CD14, and CCL2 were also increased in mice developing late flares. These analyses point to the role of macrophages as an important contributor to the pathogenesis of immune mediated nephritis, and further support the therapeutic potential of BTK inhibition in this disease and related conditions.

Keywords: BTK; BTK inhibitors; Immune cell infiltration; Lupus nephritis.

A one-step triplex RT-PCR method was characterised that allows rapid, strain-specific detection of potato virus Y (PVY) occurring on potato: PVY(N), PVY(O), PVY(NTN) (recombinant isolates), PVY(N)Wi and PVY(C). Three specific primer pairs were designed on aligned PVY sequences available from genomic data banks. The specificity of the selected primers was first examined by simplex RT-PCR with a large number of PVY reference isolates. Two fragments of 0.44 and 1.11kb were amplified for PVY(N) and non-recombinant PVY(NTN) isolates, two fragments of 0.53 and 0.66kb for PVY(O) isolates, a single fragment of 0.44kb for recombinant PVY(NTN) isolates, a 0.66kb fragment for PVY(C) isolates and a 0.53kb fragment for PVY(N)Wi isolate. The primers were then combined in a one-step triplex RT-PCR reaction, optimised stepwise and validated with the reference isolates. The great similarity between the genomes of PVY(N) and non-recombinant PVY(NTN) prevented their differentiation using this method. No fragments were amplified with samples infected by non-related potato viruses, as well as with samples from healthy tobacco and potato plants. The one-step triplex RT-PCR described here fastens specific detection of PVY strains that are otherwise only distinguishable by combined serological and biological assays.

Twenty potato virus Y (PVY) isolates were characterized. They represented two strains only, PVY(O) (three isolates) and PVY(N) (17 isolates). However, application of serological and molecular genetic methods led to a more complicated characterization. For example, five isolates induced necrotic symptoms on tobacco plants typical of PVY(N), despite reacting as PVY(O) serologically. Moreover, the PVY isolates were not identical according to molecular genetic properties. Typical PVY(NTN) PCR products were observed for 14 isolates, but five of them (Hr 220-5, Hr 387-7, Nord 242, Syn1Scot, and 41-97) did not produce potato tuber necrotic symptoms in infected cultivars. An immunocapture reverse transcription-polymerase chain reaction (RT-PCR) probing was developed using a set of 24 primer pairs derived from eight regions of the PVY genome. Using this method, five out of seven PVY(NTN) isolates including the Czech standard PVY(NTN) from the potato cv. Nicola were found to be identical. However, two PVY(NTN) isolates and all the other probed PVY samples showed unique patterns, suggesting specific differences at the nucleotide level. This method enabled specific identification of individual isolates variability even within different PVY strains.

The year 1958, when DNA was first made in a test tube, marked the birth of modern biotechnology. DNA has now developed into an important technology that makes a key contribution in various sectors such as agriculture, environment and cleansing, and it has triggered a veritable boom in medicine. Today, biologics account for more than 60% of newly approved drugs. They are efficient, save time and money and have few or minimal side effects, fuelling the appetite of big pharma to take over biotech companies. Where will the journey lead?

For the first time in the 40-year history of the National Atmospheric Deposition Program/National Trends Network (NADP/NTN), a unique urban-to-rural transect of wet deposition monitoring stations was operated as part of the NTN in 2017 to quantify reactive inorganic nitrogen wet deposition for adjacent urban and rural, montane regions. The transect of NADP stations (sites) was used to collect continuous precipitation depth and weekly wet-deposition samples in the Denver - Boulder, Colorado, urban corridor. Gradients in reactive inorganic nitrogen (Nr) concentrations and wet deposition were identified along the transect, which included Rocky Mountain National Park. Back trajectory modeling and stable isotopes suggested contribution of agricultural ammonia (NH₃) to urban Nr wet deposition in Denver, but apportionment of wet-deposited Nr to agricultural versus urban mobile sources was not possible for this study. The results demonstrate the importance of multiple monitoring sites across an urban area in defining fine-scale geographic patterns in atmospheric deposition and its sources. Data from new sites located within 50 km of the urban area demonstrate that the urban influence does not extend as far as the inverse distance weighting would have suggested without such empirical monitoring data. It is important to determine the radius of influence of urban emissions and associated deposition on the interpolated deposition raster, which is constrained by a paucity of monitoring sites east of Denver.

The velocity distributions in the clearance gap of the Kyoto-NTN biocentrifugal ventricular assist device model were measured by laser Doppler velocimetry (LDV) at three inlet flow conditions, namely operating, fully opened, and fully closed conditions. The results obtained have a similar trend as in the earlier measurements using air as medium and the hot-wire probe, a washout mechanism that is a segment of fluids in the gap situated from theta = 60 degrees to 220 degrees, has a larger radial velocity component flowing toward the eye of the pump, as compared to other regions in the gap where the tangential velocity component is dominant. It is essential to have a good washout for the leakage flow through the clearance gap between the stationary casing and the impeller of the pump so that the blood will not keep on circulating in the gap but is washed out to the eye in order to reduce the chances of being destroyed. Although the detailed velocity distributions are not the same, this should be due to the minor fabrication differences between two pump models. The current noninvasive LDV measurements should have a better representation of the actual flow field than the earlier studies due to the blood analog being used as the flow medium. Furthermore, as compared to the methods used in the earlier studies, there is basically no modification of the pump geometry in the present measurement.

Objective: Cloning and secretory expression of an amidase from Kluyvera cryocrescens and characterization of its potential in preparation of chiral amino acids.

Results: An amidase belonging to the Ntn-hydrolase superfamily was identified from Kluyvera cryocrescens ZJB-17005 (Kc-Ami). The maximum activity of Kc-Ami was observed at pH 8.5 and 55 °C. Remarkably, Kc-Ami showed an excellent enantioselectivity (99% ee) using rac-4-(hydroxy(methyl)phosphoryl)-2-(2-phenylacetamido) butanoic acid as substrate. Kc-Ami remained stable at pH 7.0-9.0 and exhibited prominent thermostability with a half-life time of 59.1, 47.4 and 20.4 h at 50, 55 and 60 °C, respectively. Kc-Ami could be appllied to synthesize chiral amino acids and its derivatives with excellent enantioselectivity (> 99% ee). The synthesized chiral amino acids could contain short or long side chain, and further the side chain could be replaced with -OH, -COOH or benzene ring.

Conclusions: Kc-Ami exhibited remarkable thermostability and excellent enantioselectivity for synthesizing chiral amino acids and its derivatives. This specific characteristic provides great potential for industrial application in preparation of chiral amino acids and its derivatives.

Keywords: Amidase; Chiral amino acid; Kluyvera cryocrescens; L-phosphinothricin; Ntn-hydrolase superfamily.

β-Aminopeptidases comprise a class of enzymes with functional and structural similarities. All members of the β-aminopeptidases described to date were isolated from bacterial sources. Uniquely, they catalyze the hydrolysis of β(3) - and/or β(2) -amino acid residues from amides and peptides that are otherwise considered proteolytically stable. Due to this unusual reactivity with β-peptide substrates, β-aminopeptidases have potential to be used as biocatalysts for β-peptide synthesis and for the resolution of enantiomerically pure β-amino acids from racemic substrate mixtures. β-Aminopeptidases are formed from an inactive precursor by posttranslational autoproteolytic cleavage, exposing the catalytic nucleophile at the N-terminus of the newly formed β-polypeptide chain. Such an activation step is a characteristic trait of enzymes of the N-terminal nucleophile (Ntn) hydrolase superfamily. However, classical Ntn hydrolases and β-aminopeptidases differ by the fold of their catalytic cores and are hence likely to originate from distinct evolutionary ancestors. In this contribution, we review the existing literature on β-aminopeptidases, including biochemical and functional studies, as well as structural investigations that recently allowed insights into the catalytic mechanisms of precursor processing and β-peptide conversion.

Acute and chronic stages of NTN are followed by infiltration of glomeruli with monocytes/macrophages having different location (lumen of capillaries in the acute stage and mesangial zone in the chronic one). TNF-alpha is one of the key factors of the NTN acute stage damaging glomerular structures and initiating production of matrix form of the main factor of the fibroblast growth and transforming growth factor beta. Accumulation of these cytokines in the matrix facilitated monocyte penetration in the mesangium zone in chronization. Interaction of the matrix-associated cytokines regulates proliferative and fibrogenic activity of the mesangial cells as well as production of TNF-alpha by monocytes/macrophages. A decrease of TNF-alpha level during a chronic stage of NTN results in a decrease of the mesangial cell ability to produce matrix-associated cytokines.