Dieback and Wilting Caused by Tomato spotted wilt virus in Arctotis × hybrida in Italy.

In winter 2012, some potted plants of African daisy (Arctotis × hybrida L., family Asteraceae) cv. Hannah, propagated by rooted stem cuttings and cultivated for commercial purposes in a greenhouse located at Albenga (Liguria region, Italy), were noticed for a rapid dieback, generalized reddening, following by an irreversible wilting. Around 130 plants on a total of 3,000 cultivated plants showed symptoms (4 to 5%). One gram of fresh leaves, each collected from three different symptomatic plants, was ground in 4 ml of cold (∼5°C) sodium phosphate 0.03 M buffer, containing 0.2% sodium diethyldithiocarbamate, 75 mg/ml of active charcoal, and traces of carborundum (600 mesh). The inoculum was rubbed on healthy indicator herbaceous plants and inoculated plants were maintained in an insect-proof greenhouse with natural illumination and temperatures of 24/18°C day/night. Healthy and buffer inoculated plants were also included in the test and used as negative control in the subsequent serological and molecular analysis. Sap-inoculated plants showed the following symptoms after 1 to 3 weeks: necrotic local lesions in Chenopodium amaranticolor and C. quinoa, yellowing and stunting following by systemic necrosis and death of the plants in tomato (Solanum lycopersicum cv. San Marzano), necrotic local lesions following by systemic necrotic patterns and leaf deformation in tobacco (Nicotiana tabacum cv. Xanthi nc.) and N. glutinosa, necrotic local lesions in petunia (Petunia × hybrida cv. Pink Beauty). No symptoms were recorded on buffer inoculated plants. Leaf samples from both symptomatic hosts and the three original symptomatic African daisy plants were tested by double-antibody sandwich-ELISA with polyclonal antisera against Cucumber mosaic virus (CMV) and tospoviruses (Tospovirus broad-spectrum, Serogroups I, II, and III, Bioreba AG, Switzerland). Positive reaction was obtained with Tospo-groups antibodies, but not with the CMV ones. Total RNA was extracted from infected leaves of African daisy with the RNeasy Plant Mini Kit (Qiagen, Valencia, CA) and subjected to reverse transcription (RT)-PCR by using the tospovirus universal primers BR60/BR65 that amplify part of the nucleocapsid protein gene (1). Target amplicons of 454 bp were produced for all samples tested. The PCR products were cloned and sequenced on both strands (one clone per amplicon cloned). The resulting sequences were 100% identical, so a single sequence was deposited in GenBank (HF913777). The sequence showed highest homology (99%) with the Tomato spotted wilt virus (TSWV) tomato isolate NJ-JN from South Korea (HM581936). The identity of the virus infecting African daisy was further confirmed by sequencing amplicons obtained by RT-PCR using primers partially covering the movement protein gene of TSWV (2). The sequence obtained (HF913776) showed the highest homology (99%) with three TSWV isolates: a tomato isolate from Spain (AY744493), a pepper isolate from South Korea (AB663306), and again the tomato NJ-JN isolate from South Korea (HM581936). To our knowledge, this is the first natural report of TSWV infecting African daisy plants. Moreover, since this ornamental is often cultivated with other flowering plants, it can act as reservoir for the virus that can infect other ornamentals and crops, considering that TSWV has a very broad host range (3). This result also represents the first finding of TSWV in the genus Arctotis, family Asteraceae, the greater botanical family of TSWV hosts (3). References: (1) M. Eiras et al. Fitopatol. Bras. 26:170, 2001. (2) M. M. Finetti et al. J. Plant Pathol. 84:145, 2002. (3) G. Parrella et al. J. Plant Pathol. 85:227. 2003.

First Record of Alfalfa mosaic virus in Teucrium fruticans in Italy.

The Teucrium genus (Lamiaceae family) contains ~300 species of evergreen and deciduous shrubs with some species widely used as ornamental plants in rock gardens. During the springs of 2010 and 2011, some plants of Teucrium fruticans L., also known as "tree germander", growing singly in pots in a Ligurian nursery (Savona Province, northern Italy), were noted for a bright yellow calico mosaic on the leaves (~1% of ~2,000 plants inspected exhibited symptoms). Preliminary electron microscope observations of leaf-dips showed semispherical to bacilliform particles, consistent with Alfamovirus and Oleavirus, in preparations obtained only from leaves of symptomatic plants. Three symptomatic and two asymptomatic plants were checked for Cucumber mosaic virus or Alfalfa mosaic virus (AMV) in protein A sandwich (PAS)-ELISA with commercial kits (Bioreba, Reinach, Switzerland) and Olive latent virus 2 (OLV2) by immunodecoration of virus particles with an OLV2 antiserum produced against an Italian OLV2 isolate. Symptomatic plants were positive only to AMV and all asymptomatic plants were negative to all viruses checked. The virus was successfully transmitted mechanically to Chenopodium amaranticolor and Ocimum basilicum that reacted, as expected for infections caused by AMV (1), with a chlorotic local lesion followed by mosaic and bright yellow mosaic, respectively. The disease was transmitted also by grafting an infected scion on healthy T. fruticans. Symptoms appeared after ~3 weeks in one plant of six grafted. AMV infection in a symptomatic grafted plant was verified by PAS-ELISA, confirming that bright yellow mosaic symptoms observed in T. fruticans were induced by an isolate of AMV. Immunocapture reverse transcription (IC-RT)-PCR assay, following the protocol described by Wetzel et al. (4), was performed on leaf extracts from one symptomatic plant using a polyclonal serum raised against a French isolate of AMV, provided by H. Lot (INRA, Station de Pathologie Végétale, Avignon, France). Specific AMV primer pair was used in the RT-PCR reactions (2). A DNA fragment of ~750 bp, covering the entire coat protein gene (CP), was obtained after IC-RT-PCR. The amplicon was gel purified with the Wizard SV Gel and PCR Clean-Up System (Promega, Madison, WI), cloned into pGEMT-easy vector (Promega) and two independent clones sequenced on both strands at MWG Biotech (Ebersberg, Germany). The consensus sequence was submitted to EMBL (No. FR854391). Pairwise comparison of the AMV-T. fruticans isolate CP sequence (named Tef-1) with those of AMV reference isolates revealed the maximum (98.0 to 97.3%) nucleotide identities with isolates belonging to subgroup I, 95.5 to 94.0% identities with subgroup IIA isolates, and 95.6% identity with the subgroup IIB isolate Tec-1 (3). Among subgroup I isolates, Tef-1 had the maximum CP nucleotide identity with the CP gene belonging to an AMV isolate identified in 2010 in Lavandula stoechas in the same geographic area, suggesting a common origin for these two viral isolates. Overall results clearly indicate that an AMV isolate was the causal agent of the calico-type mosaic observed in T. fruticans. To our knowledge, this is the first report of T. fruticans as a natural host of AMV. References: (1) G. Marchoux et al. Page 163 in: Virus des Solanacées. Quae éditions, Versailles, 2008. (2) G. +Parrella et al. Arch. Virol. 145:2659, 2000. (3) G. Parrella et al. Arch. Virol. 156:1049, 2011. (4) T. Wetzel et al. J. Virol. Methods 39:27, 1992.

Effects of ischemia on the human pancreas.

BACKGROUND & AIMS: Patients undergoing cardiac surgery frequently have pancreatic damage; several factors, including tissue hypoperfusion, have been implicated. The aim of this study is to better understand the effects of decreased blood supply on human pancreas. METHODS: Twenty-one consecutive patients undergoing surgery for thoracic or thoracoabdominal aortic aneurysm were studied. During surgery, the descending thoracic aorta was cross-clamped for a mean of 44 minutes (range, 15-85 minutes). Effects of the resultant pancreatic ischemia were assessed by examining the patients daily for evidence of acute pancreatitis for at least 20 days after surgery and by determining serum concentrations of amylase, pancreatic isoamylase, and lipase before clamping the aorta and at varying intervals after its release (1, 2, and 6 hours during the first day and once daily for the following 6 days). RESULTS: One patient died of acute necrotizing pancreatitis. None of the others had symptoms of pancreatitis postoperatively, but all showed a significant increase in serum pancreatic enzyme concentrations soon after declamping; this peaked about 24 hours later and persisted for all 7 days of the study. A significant relationship was found between the increase in pancreatic enzyme concentrations and the aortic clamping time. CONCLUSIONS: Acinar cell injury is a constant, rapidly appearing consequence of severe pancreatic ischemia, even that of brief duration; acinar cell injury is usually subclinical but may also present as severe acute pancreatitis.