The first detection of Botryosphaeria sinensis on white lupine

Fungal strain MF KP-12.1, which is similar in morphological and cultural characteristics to Botryosphaeria fungi, was isolated from the seeds of white lupine variety Dega grown in Oryol region in 2018. Phylogenetic analysis of the large rRNA subunit (LSU), elongation factor-1α (TEF) and internal transcribed spacer (ITS) fragments was used to accurately identify the isolated strain as B. sinensis. When 2-week-old white lupine plants were treated under laboratory conditions with mycelial suspension of B. sinensis MF KP-12.1, plant damage reached 67 % on day 3, and 100 % plant death was observed on day 10. Pathogenicity of B. sinensis MF KP-12.1 was confirmed by re-isolation of the strain from damaged plant tissue. This is the first detection of B. sinensis on white lupine, which has not been previously reported as a host for this fungus. It is also the first detection of B. sinensis in Russia.

Select Porcine Elongation Factor 1α Sequences Mediate Stable High-Level and Upregulated Expression of Heterologous Genes in Porcine Cells in Response to Primate Serum

Genetically engineered (GE) pigs with various combinations of genetic profiles have been developed using heterologous promoters. This study aimed to identify autologous promoters for high and ubiquitous expression of xenotransplantation relevant genes in GE pigs. A 1.4 kb upstream regulatory sequence of porcine elongation factor 1α (pEF1α) gene was selected and isolated for use as a promoter. Activity of the pEF1α promoter was subsequently compared with that of the cytomegalovirus (CMV) promoter, CMV enhancer/chicken β-actin (CAG) promoter, and human EF1α (hEF1α) promoter in different types of pig-derived cells. Comparative analysis of luciferase and mutant human leukocyte antigen class E-F2A-β-2 microglobulin (HLA-E) expression driven by pEF1α, CMV, CAG, and hEF1α promoters revealed the pEF1α promoter mediated comparable expression levels with those of the CAG promoter in porcine ear skin fibroblasts (PEFs) and porcine kidney-15 (PK-15) cells, but lower than those of the CAG promoter in porcine aortic endothelial cells (PAECs). The pEF1α promoter provided long-term stable HLA-E expression in PEFs, but the CAG promoter failed to sustain those levels of expression. For xenogeneic serum-induced cytotoxicity assays, the cells were cultured for several hours in growth medium supplemented with primate serum. Notably, the pEF1α promoter induced significant increases in luciferase and HLA-E expression in response to primate serum in PAECs compared with those driven by the CAG promoter, suggesting the pEF1α promoter could regulate temporal expression of heterologous genes under xenogeneic-cytotoxic conditions. These results suggest the pEF1α promoter may be valuable for development of GE pigs spatiotemporally and stably expressing immunomodulatory genes for xenotransplantation.

The molecular systematics and diversification of a taxonomically unstable group of Asian cicada tribes related to Cicadini Latreille, 1802 (Hemiptera : Cicadidae)

The cicadas (Hemiptera: Cicadidae) related to tribe Cicadini exhibit some of the most remarkable phenotypes in the family, with many genera possessing striking colour patterns and unusual morphological features. This largely Asian group of 13 tribes has proven challenging for cicada taxonomists, in part because of likely convergent evolution or losses of these phenotypes. We present the first focused molecular phylogeny of this clade, including ~60 described genera. The genetic dataset contains 839 ingroup-informative sites (out of 2575) from mitochondrial cytochromec oxidase subunitI, nuclear elongation factor-1α, and nuclear acetyltransferase. We use Bayesian and maximum likelihood trees to test recent changes in tribe- and subtribe-level classification, and we reconstruct ancestral character states for potentially convergent traits influencing tribe descriptions. We use fossil and molecular clock calibrations to estimate the temporal and geographic context of the radiation. The tribes Gaeanini, Leptopsaltriini, Platypleurini, Psithyristriini, and Tosenini appear polyphyletic and in need of revision, in part because of convergent evolution of opaque wings and multiple convergent gains or losses of abdominal tubercles. Kalabita Moulton, 1923 is transferred from Platypleurini to Leptopsaltriini. Vittagaeana gen. nov. is established for Vittagaeana paviei comb. nov. and Vittagaeana dives comb. nov., formerly in Tosena. Sinosenini syn. nov. is synonymised with Dundubiina. Ayuthiini trib. nov. is established with two new subtribes for Ayuthia Distant, 1919 and Distantalna Boulard, 2009, formerly in Tosenini. For the earliest split in the tree, one common ancestor appears to have been Indian + Asian in geographic distribution and the other Asian. We estimate that the radiation began in the middle Cenozoic Era, possibly as recently as the early Miocene. The recent and steady pattern of diversification suggests that refinement of tribe diagnoses will prove challenging. http://zoobank.org:urn:lsid:zoobank.org:pub:5A6C16F4-5269-453B-BA5C-B29C3394683A

Cryphonectriaceae associated with rust-infected Syzygium jambos in Hawaii

Syzygium jambos (Myrtales, Myrtaceae) trees in Hawaii are severely affected by a rust disease caused by Austropuccinia psidii (Pucciniales, Sphaerophragmiaceae), but they are commonly co-infected with species of Cryphonectriaceae (Diaporthales). In this study, S. jambos and other trees in the Myrtales were examined on three Hawaiian Islands for the presence of Cryphonectriaceae. Bark samples with fruiting bodies were collected from infected trees and fungi were isolated directly from these structures. Pure cultures were produced and the fungi were identified using DNA sequence data for the internal transcribed spacer (ITS) region, part of the β-tubulin (BT1) gene and the transcription elongation factor-1α (TEF1) gene. Five species in three genera of Cryphonectriaceae were identified from Myrtaceae tree samples. These included Chrysoporthe deuterocubensis, Microthia havanensis and three previously-unknown taxa described here as Celoporthe hauoliensis sp. nov., Cel. hawaiiensis sp. nov. and Cel. paradisiaca sp. nov. Representative isolates of Cel. hauoliensis, Cel. hawaiiensis, Cel. paradisiaca, Chr. deuterocubensis and Mic. havanensis were used in artificial inoculation studies to consider their pathogenicity on S. jambos. Celoporthe hawaiiensis, Cel. paradisiaca and Chr. deuterocubensis produced lesions on young S. jambos trees in inoculation trials, suggesting that, together with A. psidii, they may contribute to the death of trees. Microsatellite markers were subsequently used to consider the diversity of Chr. deuterocubensis on the Islands and thus to gain insights into its possible origin in Hawaii. Isolates of this important Myrtaceae and particularly Eucalyptus pathogen were found to be clonal. This provides evidence that Chr. deuterocubensis was introduced to the Hawaiian Islands as a single introduction, from a currently unknown source.

Molecular and alkaloid characterization of Claviceps purpurea sensu lato from grass seed production areas of the U.S. Pacific Northwest

Ergot, caused by Claviceps purpurea sensu lato, is an economically important seed replacement disease of Kentucky bluegrass (Poa pratensis) and perennial ryegrass (Lolium perenne) seed crops. Claviceps purpurea sensu stricto is considered the primary Claviceps species responsible, but genetic diversity and cryptic species within C. purpurea sensu lato have previously been reported. Fifty-six C. purpurea sensu lato isolates collected from P. pratensis (n=21) and L. perenne (n=35) in Oregon and Washington between 2010 and 2014 were characterized using RAPD, partial ITS, β-tubulin and elongation factor-1α sequences, conidial size, and ergot alkaloid chemotype. Based on RAPD analysis, 7 isolates from P. pratensis and 33 isolates from L. perenne collected in Oregon corresponded to C. purpurea sensu stricto, while 13 isolates collected from P. pratensis in Washington and Oregon were identified as C. humidiphila. Partial ITS, β-tubulin, and elongation factor-1α sequences identified 10 isolates from P. pratensis as C. humidiphila, while seven isolates from P. pratensis and 33 isolates from L. perenne were identified as C. purpurea sensu stricto. Several isolates generated ambiguous RAPD bands and/or sequences that prevented identification. Ergot alkaloid chemotype profiling found that ergocornine and its epimer were predominant in sclerotia from P. pratensis, whereas ergotamine and its epimer were found in highest abundance in sclerotia from L. perenne. This study confirms the presence of the C. purpurea sensu lato species complex in the U.S. Pacific Northwest and suggests that more research is needed to characterize and mitigate Claviceps spp. infection of grass seed crops in North America.

Evaluation of Cysteine Protease C of Leishmania donovani in Comparison with Glycoprotein 63 and Elongation Factor 1α for Diagnosis of Human Visceral Leishmaniasis and for Posttreatment Follow-Up Response

ABSTRACT Visceral leishmaniasis (VL) is a threat in many developing countries. Much effort has been put to eliminating this disease, for which serodiagnosis remains the mainstay for VL control programs. New and improved antigens as diagnostic candidates are required, though, as the available antigens fail to demonstrate equal optimum performance in all areas of endemicity. Moreover, these diagnoses are dependent on invasive serum sampling. In the current study, we cloned and expressed Leishmania donovani cysteine protease C (CPC) and evaluated its diagnostic and test-of-cure possibilities by detecting the antibody levels in human serum and urine through ELISA and immunoblot assays. Two immunodominant antigens, recombinant glycoprotein 63 (GP63) and elongation factor 1α (EF1α), identified earlier by our group, were also assessed by employing human serum and urine samples. Of these three antigens in ELISAs, CPC demonstrated the highest sensitivities of 98.15% and 96% positive testing in serum and urine of VL patients, respectively. Moreover, CPC yielded 100% specificity with serum and urine of nonendemic healthy controls compared to GP63 and EF1α. Urine samples were found to be more specific than serum for distinguishing endemic healthy controls and other diseases by means of all three antigens. In all cases, CPC gave the most promising results. Unlike serum, urine tests demonstrated a significant decrease in antibody levels for CPC, GP63, and EF1α after 6 months of treatment. The diagnostic and test-of-cure performances of CPC in the immunoblot assay were found to be better than those of GP63 and EF1α. In conclusion, CPC, followed by GP63 and EF1α, may be utilized as candidates for diagnosis of VL and to assess treatment response.

Gnomoniopsis chinensis (Gnomoniaceae, Diaporthales), a new fungus causing canker of Chinese chestnut in Hebei Province, China

Chinese chestnut (Castanea mollissima) is an important crop tree species in China. However, branch canker and fruit rot are two kinds of severe diseases, which weaken the host and decrease chestnut production. During our investigations into chestnut diseases in China, several fungi have been confirmed as casual agents in previous studies, namely Aurantiosacculus castaneae, Cryphonectria neoparasitica, Cry. parasitica, Endothia chinensis and Gnomoniopsis daii. In this study, a new canker pathogen is introduced based on morphology, phylogeny and pathogenicity. Typical Gnomoniopsis canker sign of wide, orange tendrils emerging from hosts’ glaucous lenticels were obvious on the diseased trees in the field. Symptomatic branches or bark on stems from different chestnut plantations were sampled and isolated, then strains were identified by comparisons of DNA sequence data for the nuclear ribosomal internal transcribed spacer (ITS), partial translation elongation factor-1α (tef1) and β-tubulin (tub2) gene regions as well as morphological features. As a result, these strains appeared different from any known Gnomoniopsis species. Hence, we propose a novel species named Gnomoniopsis chinensis. Pathogenicity was further tested using the ex-type strain (CFCC 52286) and another strain (CFCC 52288) on both detached branches and 3-year-old chestnut seedlings. The inoculation results showed that Gnomoniopsis chinensis is mildly pathogenic to Chinese chestnut. However, further studies are required to confirm its pathogenicity to the other cultivated Castanea species in America, Europe and Japan.

Specific Detection and Identification of Fusarium graminearum Sensu Stricto Using a PCR-RFLP Tool and Specific Primers Targeting the Translational Elongation Factor 1α Gene

Fusarium graminearum is a toxigenic plant pathogen that causes Fusarium head blight (FHB) disease on cereal crops. It has recently shown to have cross-pathogenicity on noncereals (i.e., Fusarium root rot [FRR] on soybean) in Canada and elsewhere. Specific detection and differentiation of this potent toxigenic, trichothecene-producing pathogen among other closely related species is extremely important for disease control and mycotoxin monitoring. Here, we designed a PCR restriction fragment length polymorphism protocol based on the DNA sequence of the translational elongation factor 1α (TEF1α) gene. A unique restriction site to the enzyme HpaII is only found in F. graminearum sensu stricto strains among different Fusarium strains in the F. graminearum species complex (FGSC) and other Fusarium spp. associated with FHB in cereals and FRR in soybean. Partial amplification of the TEF1α gene with newly designed primers mh1/mh2 generated a 459-bp PCR fragment. Restriction digestion of the generated fragments with the HpaII enzyme generated a unique restriction pattern that can rapidly and accurately differentiate F. graminearum sensu stricto among all other Fusarium spp. A primer pair (FgssF/FgssR) specific to F. graminearum sensu stricto also was designed and can distinguish F. graminearum sensu stricto from all other Fusarium spp. in the FGSC and other closely related Fusarium spp. involved in FHB and FRR. This finding will be very useful for the specific detection of F. graminearum sensu stricto for diagnostic purposes as well as for the accurate detection of this pathogen in breeding and other research purposes.