Establishment and characterization of American elm cell suspension cultures

Cell suspension cultures of Dutch elm disease (DED)-tolerant and DED-susceptible American elms clones have been established and characterized as prerequisites for contrasts of cellular responses to pathogen-derived elicitors. Characteristics of cultured elm cell growth were monitored by A₇₀₀ and media conductivity. Combined cell growth data for all experiments within a genotype showed relatively low variances and between-genotype contrasts during repeated passages showed no significant differences. Subculturing exponentially growing cells at 8–14 day intervals, within readily measured parameters of media conductivity (4.95–4.2 mmhos) and cell concentration (≥ 1.4 A₇₀₀), consistently resulted in repeatable profiles of elm cell growth and minimized lag phase. Culture cells were essentially homogeneous after 5 subculture passages and their overall appearance was stable. We conclude that the described procedure resulted in consistent cultures suitable for elicitor treatment experiments. This revised version was published online in June 2006 with corrections to the Cover Date.     

An unusually long Rift valley fever inter-epizootic period in Zambia: Evidence for enzootic virus circulation and risk for disease outbreak

Rift valley fever (RVF) is a mosquito-borne disease of animals and humans. Although RVF outbreaks are usually reported at 5-15-year intervals in sub-Saharan Africa, Zambia has experienced an unusually long inter-epizootic/-epidemic period of more than three decades. However, serological evidence of RVF virus (RVFV) infection in domestic ruminants during this period underscores the need for comprehensive investigation of the mechanisms of virus perpetuation and disease emergence. Mosquitoes (n = 16,778) captured from eight of the ten provinces of Zambia between April 2014 and May 2019 were pooled (n = 961) and screened for RVFV genome by a pan-phlebo RT-PCR assay. Aedes mosquito pools (n = 85) were further screened by nested RT-PCR assay. Sera from sheep (n = 13), goats (n = 259) and wild ungulates (n = 285) were screened for RVFV antibodies by ELISA while genome detection in pooled sera (n = 276) from domestic (n = 248) and wild ungulates (n = 37) was performed by real-time RT-PCR assay. To examine the association between the long inter-epizootic period and climatic variables, we examined El Niño-Southern Oscillation indices, precipitation anomalies, and normalized difference vegetation index. We then derived RVF risk maps by exploring climatic variables that would favor emergence of primary RVFV vectors. While no RVFV genome could be detected in pooled mosquito and serum samples, seroprevalence was significantly high (OR = 8.13, 95% CI [4.63–14.25]) in wild ungulates (33.7%; 96/285) compared to domestic ruminants (5.6%; 16/272). Retrospective analysis of RVF epizootics in Zambia showed a positive correlation between anomalous precipitation (La Niña) and disease emergence. On risk mapping, whilst northern and eastern parts of the country were at high risk, domestic ruminant population density was low (< 21 animals/km²) in these areas compared to low risk areas (>21 animals/km²). Besides evidence of silent circulation of RVFV and the risk of disease emergence in some areas, wildlife may play a role in the maintenance of RVFV in Zambia.     

Chemical analysis and study of immunoenhancing and antioxidant property of a glucan isolated from an alkaline extract of a somatic hybrid mushroom of Pleurotus florida and Calocybe indica variety APK2

A water-soluble glucan isolated from an alkaline extract of fruit bodies of a somatic hybrid mushroom PCH9FB of Pleurotus florida and Calocybe indica var. APK2 strains showed antioxidant properties with immune activation of macrophage, splenocyte, and thymocyte. On the basis of acid hydrolysis, methylation, periodate oxidation and NMR studies ((1)H, (13)C, DEPT-135, DQF-COSY, TOCSY, NOESY, ROESY, HMQC, and HMBC) the structure of the repeating unit of the glucan was established as: [structure: see text].     

Adenylate cyclase 5 is required for melanophore and male pattern development in the guppy (Poecilia reticulata)

Guppies (Poecilia reticulata) are colorful fish that have attracted the attention of pigmentation researchers for almost a century. Here, we report that the blond phenotype of the guppy is caused by a spontaneous mutation in the guppy ortholog of adenylate cyclase 5 (adcy5). Using double digest restriction site‐associated DNA sequencing (ddRADseq) and quantitative trait locus (QTL) mapping, we linked the blond phenotype to a candidate region of 118 kb, in which we subsequently identified a 2‐bp deletion in adcy5 that alters splicing and leads to a premature stop codon. We show that adcy5, which affects life span and melanoma growth in mouse, is required for melanophore development and formation of male orange pigmentation traits in the guppy. We find that some components of the male orange pattern are particularly sensitive to loss of Adcy5 function. Our work thus reveals a function for Adcy5 in patterning of fish color ornaments.     

Muscle‐specific Cand2 is translationally upregulated by mTORC1 and promotes adverse cardiac remodeling

The mechanistic target of rapamycin (mTOR) promotes pathological remodeling in the heart by activating ribosomal biogenesis and mRNA translation. Inhibition of mTOR in cardiomyocytes is protective; however, a detailed role of mTOR in translational regulation of specific mRNA networks in the diseased heart is unknown. We performed cardiomyocyte genome‐wide sequencing to define mTOR‐dependent gene expression control at the level of mRNA translation. We identify the muscle‐specific protein Cullin‐associated NEDD8‐dissociated protein 2 (Cand2) as a translationally upregulated gene, dependent on the activity of mTOR. Deletion of Cand2 protects the myocardium against pathological remodeling. Mechanistically, we show that Cand2 links mTOR signaling to pathological cell growth by increasing Grk5 protein expression. Our data suggest that cell‐type‐specific targeting of mTOR might have therapeutic value against pathological cardiac remodeling.     

Ozone sensitivity in hybrid poplar correlates with insensitivity to both salicylic acid and jasmonic acid. The role of programmed cell death in lesion formation

Our earlier studies demonstrated that the ozone-sensitive hybrid poplar clone NE-388 displays an attenuated level of ozone-, wound-, and phytopathogen-induced defense gene expression. To determine if this reduced gene activation involves signal transduction pathways dependent on salicylic acid (SA) and/or jasmonic acid (JA), we compared the responses of NE-388 and an ozone-tolerant clone, NE-245, to these signal molecules. JA levels increased in both clones in response to ozone, but only minimal increases in SA levels were measured for either clone. Treatment with SA and methyl jasmonate induced defense gene expression only in NE-245, indicating that NE-388 is insensitive to these signal molecules. DNA fragmentation, an indicator of programmed cell death (PCD), was detected in NE-245 treated with either ozone or an avirulent phytopathogen, but was not detected in NE-388. We conclude that these clones undergo two distinct mechanisms of ozone-induced lesion formation. In NE-388, lesions appear to be due to toxic cell death resulting from a limited ability to perceive and subsequently activate SA- and/or JA-mediated antioxidant defense responses. In NE-245, SA-dependent PCD precedes lesion formation via a process related to the PCD pathway activated by phytopathogenic bacteria. These results support the hypothesis that ozone triggers a hypersensitive response.     

Rapamycin inhibition of CFA-induced lymphangiogenesis in PLN is independent of mast cells

Elucidation of the events responsible for the interaction between lymphatic endothelial cells (LECs) and mast cells (MCs) may prove to be a valuable source for controlling lymphangiogenesis. In the present study, we compared immunohistochemical and RT-PCR findings of the popliteal lymph node (PLN) and footpad skin in C57BL/6J and WBB6F1 mice, the MC-deficient strain. The results indicated that MCs play certain role in complete Freund’s adjuvant-induced intranodal lymphangiogenesis. VEGF-A, VEGFR-2 and TNF-α were crucial factors in lymphangiogenesis both in the PLN and skin. Moreover, the in vivo administration of the specific mTOR inhibitor, rapamycin inhibited lymphangiogenesis independent of MCs in PLN rather than in the skin. Further study on anti-lymphangiogenic effect will contribute to our understanding of LEC and MC modulation in pathological lymphangiogenesis.     

Development of oxidovanadium and oxido-peroxido vanadium-based artificial DNA nucleases via multi spectroscopic investigations and theoretical simulation of DNA binding

Benzimidazole is a neutral ligand which is often used to synthesize bioactive compounds. Two transition metal benzimidazole-based complexes, namely, vanadium (IV) dioxido complex (complex 1) and vanadium (V) oxido-peroxido complex (complex 2) with tridentate benzimidazole ligand, 2,6-di (1H-benzo[d]imidazol-2-yl) pyridine (Byim) have been designed with the intention of developing potential DNA nuclease. Different studies involving biochemical and biophysical techniques along with molecular docking suggest that both the complexes interact with DNA, while the mode of binding is intercalation. The complexes were further used for DNA cleavage activity. Both of them were found to have substantial DNA nuclease activity, but complex 2 was more potent than complex 1 in exhibiting such activity.