Feeding by Leucopis argenticollis and Leucopis piniperda (Diptera: Chamaemyiidae) from the western USA on Adelges tsugae (Hemiptera: Adelgidae) in the eastern USA.

Leucopis argenticollis (Zetterstedt) and Leucopis piniperda (Malloch) are known to feed on the lineage of Adelges tsugae Annand that is native to western North America, but it is not known if they will survive on the lineage that was introduced from Japan to the eastern USA. In 2014, western Leucopis spp. larvae were brought to the laboratory and placed on A. tsugae collected in either Washington (North American A. tsugae lineage) or Connecticut (Japanese lineage). There were no significant differences in survival or developmental times between flies reared on the two different adelgid lineages. In 2015 and 2016, western Leucopis spp. adults were released at two different densities onto enclosed branches of A. tsugae infested eastern hemlock (Tsuga canadensis (L.) Carr.) in Tennessee and New York. Cages were recovered and their contents examined 4 weeks after release at each location. Leucopis spp. larvae and puparia of the F1 generation were recovered at both release locations and adults of the F1 generation were collected at the Tennessee location. The number of Leucopis spp. offspring collected increased with increasing adelgid density, but did not differ by the number of adult flies released. Flies recovered from cages and flies collected from the source colony were identified as L.argenticollis and L. piniperda using DNA barcoding. These results demonstrate that Leucopis spp. from the Pacific Northwest are capable of feeding and developing to the adult stage on A. tsugae in the eastern USA and they are able to tolerate environmental conditions during late spring and early summer at the southern and northern extent of the area invaded by A. tsugae in the eastern USA.

Phytosanitation Methods Influence Posttreatment Colonization of Juglans nigra Logs by Pityophthorus juglandis (Coleoptera: Curculionidae: Scolytinae).

Several North American walnut species (Juglans spp.) are threatened by thousand cankers disease which is caused by the walnut twig beetle (Pityophthorus juglandis Blackman) and its associated fungal plant pathogen, Geosmithia morbida M. Kolarík, E. Freeland, C. Utley and N. Tisserat sp. nov. Spread of this disease may occur via movement of infested black walnut (Juglans nigra L.) wood. This study evaluated the ability of P. juglandis to colonize J. nigra wood previously treated with various phytosanitation methods. Steam-heated and methyl bromide-fumigated J. nigra logs, as well as kiln-dried natural wane J. nigra lumber (with and without bark) were subsequently exposed to P. juglandis colonization pressure in two exposure scenarios. Following a pheromone-mediated, high-pressure scenario in the canopy of infested trees, beetles readily colonized the bark of steam-heated and methyl bromide-fumigated logs, and were also recovered from kiln-dried lumber on which a thin strip of bark was retained. In the simulated lumberyard exposure experiment, during which samples were exposed to lower P. juglandis populations, beetles were again recovered from bark-on steam-heated logs, but were not recovered from kiln-dried bark-on lumber. These data suggest logs and bark-on lumber treated with phytosanitation methods should not be subsequently exposed to P. juglandis populations. Further beetle exclusion efforts for phytosanitized, bark-on walnut wood products transported out of quarantined areas may be necessary to ensure that these products do not serve as a pathway for the spread of P. juglandis and thousand cankers disease.

Ipsenol, Ipsdienol, Ethanol, and α-Pinene: Trap Lure Blend for Cerambycidae and Buprestidae (Coleoptera) in Pine Forests of Eastern North America.

In 2007-2008, we examined the flight responses of wood-boring beetles (Coleoptera: Cerambycidae and Buprestidae) to multiple-funnel traps baited with the pine volatiles, ethanol, and α-pinene [85% (-)], and the bark beetle pheromones, racemic ipsenol and racemic ipsdienol. Experiments were conducted in mature pine stands in Canada (Ontario and New Brunswick) and the United States (Arkansas, Florida, Michigan, New Hampshire, North Carolina, Ohio, Tennessee, and Wisconsin). At each location, traps were deployed in 10 replicate blocks of four traps per block. The trap treatments were: 1) blank control; 2) ipsenol and ipsdienol; 3) ethanol and α-pinene; and 4) a quaternary blend of ipsenol, ipsdienol, ethanol, and α-pinene. Traps baited with the quaternary blend caught the greatest numbers of Acanthocinus nodosus (F.), Acanthocinus obsoletus (Olivier), Acmaeops proteus (Kirby), Astylopsis sexguttata (Say), Rhagium inquisitor (L.) (Cerambycidae), and Buprestis lineata (F.) (Buprestidae). Traps baited with ethanol and α-pinene caught the greatest numbers of Arhopalus rusticus (LeConte), Asemum striatum (L.), Tetropium spp., Xylotrechus sagittatus (Germar) (Cerambycidae), and Buprestis maculipennis Gory (Buprestidae) with minimal interruption by ipsenol and ipsdienol. Our results suggest that multiple-funnel traps baited with the quaternary lure blend of ipsenol, ipsdienol, ethanol, and α-pinene are effective for trapping various species of wood-boring beetles in pine forests of eastern North America, and may have utility in detection programs for adventive species in North America and overseas.

Efficacy of heat treatment for the thousand cankers disease vector and pathogen in small black walnut logs.

Thousand cankers disease, caused by the walnut twig beetle (Pityophthorus juglandis Blackman) and an associated fungal pathogen (Geosmithia morbida M. Kolarík, E. Freeland, C. Utley, and N. Tisserat), threatens the health and commercial use of eastern black walnut (Juglans nigra L.), one of the most economically valuable tree species in the United States. Effective phytosanitary measures are needed to reduce the possibility of spreading this insect and pathogen through wood movement. This study evaluated the efficacy of heat treatments and debarking to eliminate P. juglandis and C. morbida in J. nigra logs 4-18 cm in diameter and 30 cm in length. Infested logs were steam heated until various outer sapwood temperatures (60, 65, and 70 degrees C in 2011; 36, 42, 48, 52, and 56 degrees C in 2012) were maintained or exceeded for 30-40 min. In 2011, all heat treatments eliminated G. morbida from the bark, but logs were insufficiently colonized by P. juglandis to draw conclusions about treatment effects on the beetle. Debarking did not ensure elimination of the pathogen from the sapwood surface. In 2012, there was a negative effect of increasing temperature on P. juglandis emergence and G. morbida recovery. G. morbida did not survive in logs exposed to treatments in which minimum temperatures were 48 degrees C or higher, and mean P. juglandis emergence decreased steadily to zero as treatment minimum temperature increased from 36 to 52 degrees C. A minimum outer sapwood temperature of 56 degrees C maintained for 40 min is effective for eliminating the thousand cankers disease vector and pathogen from walnut logs, and the current heat treatment schedule for the emerald ash borer (60 degrees C core temperature for 60 min) is more than adequate for treating P. juglandis and G. morbida in walnut firewood.

Effect of tree species and end seal on attractiveness and utility of cut bolts to the redbay ambrosia beetle and granulate ambrosia beetle (coleoptera: Curculionidae: Scolytinae).

The redbay ambrosia beetle, Xyleborus glabratus Eichhoff, is a non-native invasive pest and vector of the fungus that causes laurel wilt disease in certain trees of the family Lauraceae. This study assessed the relative attractiveness and suitability of cut bolts of several tree species to X. glabratus. In 2009, female X. glabratus were equally attracted to traps baited with swampbay (Persea palustris (Rafinesque) Sargent) and camphortree (Cinnamomum camphora (L.) J. Presl), which were more attractive than avocado (Persea americana Miller), lancewood (Ocotea coriacea (Swartz) Britton), and sweetbay (Magnolia virginiana L.). These species were more attractive than loblolly bay (Gordonia lasianthus (L.) J. Ellis). X. glabratus entrance hole density and emergence from caged bolts were highest on swampbay and camphortree. In 2010, swampbay was significantly more attractive to X. glabratus than sassafras (Sassafras albidum (Nuttall) Nees), yellow poplar (Liriodendron tulipifera L.), and eastern redbud (Cercis canadensis L.). Sassafras bolts end sealed with a liquid wax-and-water emulsion were more attractive to X. glabratus than end-sealed bolts of yellow poplar and redbud. Relative to unsealed bolts, end seal decreased X. glabratus entrance hole density on swampbay and decreased granulate ambrosia beetle (Xylosandrus crassiusculus (Motschulsky)) trap catch, entrance hole density, and adult emergence from swampbay. X. crassiusculus was not attracted to sassafras, yellow poplar, and redbud and was not more attracted to manuka oil than to unbaited traps. Sassafras was more attractive to X. glabratus than previously reported and supported reproducing populations of the insect. End sealing bolts with a wax-and-water emulsion may not be optimal for attracting and rearing ambrosia beetles in small logs.

Shoot Blight Caused by Sirococcus tsugae on Eastern Hemlock (Tsuga canadensis) in Georgia.

Eastern hemlock (Tsuga canadensis) is an ecologically and economically important conifer from the north-central United States to the east coast of North America to the southern Appalachian Mountains. In early spring 2010, blighted shoot tips of eastern hemlock were observed at widely separated locations in the Chattahoochee National Forest in north Georgia. Damage did not appear to be directly related to hemlock woolly adelgid (Adelges tsugae) activity, which was sporadic or absent in some areas where symptoms were observed. A preliminary survey in March 2010 revealed that incidence of blighted shoots on individual trees varied, but was as high as 70%. Stems of shoots produced the previous year were frequently necrotic, had lost needles, and bore pycnidia with hyaline, two-celled conidia consistent with those of Sirococcus tsugae (2,3). Later in the spring and summer, shoots of the current year's growth became blighted, with sporulation of S. tsugae also on dead and dying needles. While S. tsugae previously has been reported on T. heterophylla, T. mertensiana, Cedrus atlantica, and C. deodara in western North America, it has only recently been reported on eastern hemlock (1), and its ability to induce shoot blight has not been proven. Pure cultures (2,3) were obtained on streptomycin-amended potato dextrose agar (PDA) and their identity was confirmed by species-specific PCR primers (4). Nuclear rDNA internal transcribed spacer sequence (554 nucleotides) also was obtained for isolate 10-05 and deposited in GenBank (Accession No. HQ256769). This sequence was found to be identical to sequences previously deposited for S. tsugae isolates. Isolate 10-05 and a second isolate (10-06) were used to inoculate potted 2-year-old eastern hemlock seedlings in a growth chamber at 20°C with a 16-h photoperiod. Conidia were collected by flooding 1-month-old colonies on PDA with sterile water. Expanding shoots on one branch of each seedling were wounded using scissors to cut the tips off needles and stems, while another branch remained nonwounded. Ten seedlings per isolate were inoculated by spraying to runoff with a suspension of 5 × 106 conidia ml-1 in sterile water, and five similarly treated control seedlings were sprayed with sterile water. Seedlings were covered with plastic bags to maintain high humidity for 4 days. Germination of conidia of each isolate incubated on water agar in this growth chamber was >80% after 24 h. Symptoms were evaluated and reisolation was attempted on streptomycin-amended PDA 2 months after inoculation. Symptoms of seedlings inoculated with either isolate included chlorotic and necrotic needle spots, browning of cut edges of needles, browning and death of needle tips and entire needles, death of stem tips with retention of dead needles, and needle loss. Symptoms of control seedlings were limited to slight browning of cut edges of needles. The fungus was reisolated from wounded shoots of 17 of 20 inoculated seedlings and nonwounded shoots of 5 of 20 inoculated seedlings and was not cultured from control seedlings. To our knowledge, this is the first report of S. tsugae in Georgia and also the first demonstration of its ability to produce symptoms that have been attributed to it on any tree species. References: (1) M. Miller-Weeks and W. Ostrofsky. USDA. Forest Service. Online publication. NA-PR-01-10, 2010. (2) A. Y. Rossman et al. For. Pathol. 38:47. (3) D. R. Smith et al. For. Pathol. 33:141, 2003. (4) D. R. Smith and G. R. Stanosz. For. Pathol. 38:156, 2008.

First Report of Laurel Wilt Disease Caused by Raffaelea lauricola on Pondspice in Florida.

Laurel wilt is a fungal vascular disease of redbay (Persea borbonia (L.) Spreng) and other plants in the family Lauraceae in the southeastern United States (1). The disease is caused by Raffaelea lauricola T. C. Harr., Fraedrich & Aghayeva, which is vectored by the exotic redbay ambrosia beetle (Xyleborus glabratus Eichhoff) (2). Pondspice (Litsea aestivalis (L.) Fern.) is an obligate wetland shrub listed as endangered in Florida and Maryland and threatened in Georgia (4). On 29 August 2008, 369 of 430 (85%) pondspice trees observed at St. Marks Pond in St. John's County, Florida were dead and/or dying (4). Stem samples were collected from plants with wilted and reddened foliage, entrance holes with boring dust characteristic of ambrosia beetle attack, and dark discoloration in the outer sapwood. Discolored stem sections were surface disinfested for 30 s in a 5% sodium hypochlorite solution and then plated onto cycloheximide streptomycin malt extract agar (1). Smooth, cream-buff, submerge hyphae with uneven margins resembling R. lauricola (2) was observed growing from all sapwood pieces. DNA was extracted from a single isolate (PL 392) and the 18s small subunit rDNA was PCR amplified and sequenced with primers NS1 and NS4 (3), resulting in a 1,026-bp amplicon. A BLASTn search showed identical homology to R. lauricola strain PL 159 (GenBank Accession No. EU257806). The 18s small subunit rDNA sequence was deposited into GenBank (FJ514097). In May 2011, a spore suspension was made by flooding a single-spore culture plate of isolate PL 392 with 2 ml of sterile water, collecting the spores by pipette, and quantification by hemacyometer to 1.5 × 106 spores/ml. Pathogenicity tests were conducted on 1 to 1.5 m tall pondspice plants. Six saplings were wounded by a 3/32-inch drill bit, with four receiving 50 µl of the spore suspension and two serving as water-inoculated controls. All plants were kept in a greenhouse under ambient temperature. Within 21 days, all fungal-inoculated saplings displayed complete canopy wilt, typical of laurel wilt. R. lauricola was later recovered from all four infected plants, completing Koch's postulates. To determine if the vector can reproduce in pondspice, infected stem sections were placed in a plastic rearing box indoors at room temperature, and both callow and mature adult female X. glabratus emerged in October and November 2008. Although laurel wilt has been previously observed on pondspice in South Carolina and Georgia (1), this is the first confirmation of the disease on pondspice in Florida and the first confirmation of the vector from stem material of this host. References: (1) S. W. Fraedrich et al. Plant Dis. 92:215, 2008. (2) T. C. Harrington et al. Mycotaxon 104:399, 2008. (3) M. A. Innis et al. PCR Protocols, A Guide to Methods and Applications. Academic Press. San Diego, CA, 1990. (4) J. A. Surdick and A. M. Jenkins. Pondspice (Litsea aestivalis) Population Status and Response to Laurel Wilt Disease in Northeast Florida. Florida Natural Areas Inventory, Tallahassee, FL, 2009.

First Occurrence of Laurel Wilt Disease Caused by Raffaelea lauricola on Redbay Trees in Mississippi.

Laurel wilt is a lethal, nonnative vascular wilt disease of redbay (Persea borbonia), sassafras (Sassafras albidum), and other trees in the Lauraceae (1,4). It is caused by a fungus (Raffaelea lauricola) and transmitted by the redbay ambrosia beetle (Xyleborus glabratus), a nonnative insect first detected in Georgia in 2002 (1,2). Since introduction of the pathogen and vector (presumably from Asia), laurel wilt has caused extensive mortality to redbays in Georgia, Florida, and South Carolina (1). In June 2009, a landowner in Gautier, MS reported dead redbay trees. Signs and symptoms were identical to those reported for laurel wilt along the Atlantic Coast (wilted, bronze red foliage, and dark gray-to-black vascular discoloration) (1). Infected trees have subsequently been confirmed in and near the Pascagoula River Basin. Size of infected redbays ranged from 5 to 20 cm (diameter at breast height). No heavily decomposed or fallen redbays were noted. Many individual specimens exhibited extensive drying of stem wood and dry, wilted, light brown foliage. This indicates that introduction to the area may have occurred within the last 3 years. X. glabratus adults were collected (30°26'44.45″N, 88°39'41.83″W) in a Lindgren funnel trap baited with phoebe and manuka oil lures. Beetle identification was confirmed by USDA-APHIS, and voucher specimens were submitted to the Smithsonian National Museum of Natural History and the Mississippi Entomological Museum. Symptomatic redbay wood chips from the same location were surface sterilized and plated on cycloheximide-streptomycin malt agar and R. lauricola was isolated. A 1,026-bp portion of 18S rDNA (GenBank No. GQ996063) was amplified by PCR and sequenced using primers NS1 and NS4. BLASTn searches revealed perfect homology to R. lauricola isolate PL 697 (GQ329704). Two isolates of R. lauricola were recovered and prepared into separate spore suspensions (1 × 108 CFU/ml). Each isolate was inoculated into two healthy redbays. The inoculated redbays were placed in a growth chamber with two water-only controls. All inoculated plants, and none of the controls, exhibited wilt symptoms and died within 20 days. R. lauricola was recovered from the discolored sapwood of the inoculated plants, completing Koch's postulates. A model prediction for the natural dispersion of X. glabratus and R. lauricola estimated that these organisms may not reach Mississippi for 10 to 15 years (3). The current detection of laurel wilt in Mississippi is substantially ahead of this estimate. Currently, no records of laurel wilt have been reported from western Georgia, all of Alabama, or the panhandle of Florida. Confirmed locations in Mississippi are in Jackson County, along the Interstate 10 corridor and the Pascagoula River drainage. Due to the relatively large extent of the infestation (~64 km2, including hundreds of infected trees) eradication is not being attempted. Surveys, remote sensing, and phylogeographic analysis are underway to delineate the extent of infestation and discover the mode of introduction. The current outbreak of laurel wilt in Mississippi is likely the result of human transport of infested wood, either from Asia as a separate, new introduction or from previously infested areas in the southeastern United States. References: (1) S. W Fraedrich et al. Plant Dis. 92:215, 2008. (2) T. C. Harrington et al. Mycotaxon 104:399, 2008. (3) F. Koch and W. Smith. Environ. Entomol. 37:442, 2008. (4) J. A. Smith et al. Plant Dis. 93:198, 2009.

First Report of Laurel Wilt Disease Caused by Raffaelea lauricola on Sassafras in Florida and South Carolina.

Laurel wilt disease, caused by Raffaelea lauricola (T.C. Harr., Fraedrich & Aghayeva sp. nov.), which is a fungal symbiont of the nonnative redbay ambrosia beetle (Xyleborus glabratus Eichhoff), has caused widespread mortality of native redbay (Persea borbonia (L.) Spreng) in Georgia, South Carolina, and Florida since 2002. The disease has been noted on other species in the Lauraceae including sassafras in Georgia (1), and more recently, on avocado and camphor in Florida (4). Since 2005, wilted shoots, branch dieback, and tree death have been observed in sassafras trees (Sassafras albidum (L.)) in Liberty, McIntosh, Chatham, Effingham, Bulloch, Evans, and Screven counties in Georgia; Bamberg, Beaufort, Charleston, Colleton, Hampton, and Orangeburg counties in South Carolina; and Putnam County in Florida. Symptomatic sassafras trees ranged from 1 to 12 m high and 2.5 to 25 cm in diameter at breast height. In contrast to red bay trees that retain wilted foliage, symptomatic sassafras defoliate rapidly as trees wilt and die. Multiple symptomatic ramets originating from a common root system have been observed. Removal of bark from stem and root sections from wilted trees revealed black-to-brownish staining in the sapwood, characteristic of laurel wilt. Wood chips from symptomatic areas of branches and roots were surface sterilized and plated on cycloheximide-streptomycin malt agar as previously described (1) and R. lauricola was routinely isolated. Small subunit (18S) sequences from rDNA were amplified by PCR and sequenced using primers NS1 and NS4 (3) for isolates from sassafras from Florida and South Carolina. BLASTn searches revealed homology to Raffaelea sp. C2203 (GenBank Accession No. EU123076, 100% similarity) described by Fraedrich et al. (1) from redbay and later named R. lauricola (2). The small subunit rDNA sequences for these isolates have been deposited into GenBank ( http://www.ncbi.nlm.nih.gov/Genbank/index.html ) and assigned Accession Nos. EU980448 (Florida) and GQ329704 (South Carolina). Koch's postulates have been completed with R. lauricola on this host previously (1). Laurel wilt on sassafras often was geographically isolated from other symptomatic hosts in Georgia and South Carolina and appears to occur on this host independently of proximity to redbay. Further studies to determine the epidemiology of laurel wilt on sassafras, potential resistance, and impact on sassafras life history and distribution are needed. Given the clonal nature of sassafras, the disease would appear to have the potential to move through roots of trees once established in a stand. References: (1) S. W Fraedrich et al. Plant Dis. 92:215, 2008. (2) T. C. Harrington et al. Mycotaxon 104:399, 2008. (3) M. A. Innis et al. PCR Protocols, A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990. (4) J. A. Smith et al. Plant Dis. 93:198, 2009.

First Report of Laurel Wilt Disease Caused by Raffaelea lauricola on Camphor in Florida and Georgia.

Laurel wilt is a recently described (1) vascular disease of redbay (Persea borbonia (L.) Spreng) and other plants in the family Lauraceae. The wilt is caused by Raffaelea lauricola, a fungus vectored by the nonnative redbay ambrosia beetle (Xyleborus glabratus Eichhoff) (1,2). Since 2003, laurel wilt has caused widespread mortality of redbay in Georgia, South Carolina, and Florida (1) and has recently been found on avocado in Florida (4). Since June of 2007, wilted shoots and branch dieback have been observed in several camphor trees (Cinnamomum camphora (L.) Sieb.) in residential areas of McIntosh and Glynn counties in Georgia and Baker County in Florida. Symptomatic camphor trees ranged from 4.5 to 12 m high and occurred in areas where redbay mortality due to laurel wilt has been frequently observed during the last 2 to 3 years. In some camphor trees, only the smaller branches (<2 cm in diameter) were wilting or dead, whereas in other trees (e.g., Baker County, Florida), the larger branches and substantial portions of the crown were also symptomatic. Rapid wilt that affects entire trees that is usually observed in redbay, has not been observed in camphor. Some camphor trees in residential areas of Jekyll Island, Georgia (Glynn County), where extensive wilt of redbay has occurred, have exhibited only localized wilt of some shoots or branches and other camphors remain asymptomatic. Removal of bark from wilted branch sections revealed black-to-brownish staining in the sapwood, characteristic of laurel wilt. Although no evidence of ambrosia beetles was observed on these samples, more extensive surveys are needed to determine the role of this vector in laurel wilt of camphor. Wood chips from symptomatic areas of branches were surface sterilized and plated on cycloheximide-streptomycin malt agar as previously described (1,4) and R. lauricola was routinely isolated. Small subunit (18S) sequences from rDNA were amplified by PCR and sequenced using primers NS1 and NS4 (3). BLASTn searches revealed homology to R. lauricola C2203 (GenBank Accession No. EU123076, 100% similarity, e-value of 0.0, and a total score of 1,886). The small subunit rDNA sequence for this isolate has been deposited into GenBank ( http://www.ncbi.nlm.nih.gov/Genbank/index.html ) and has been assigned Accession No. EU 853303. The presence of laurel wilt in camphor provides an opportunity to understand the pathogen distribution and possible resistance mechanisms in this host, which could have implications for efforts to remediate the impacts of the disease in redbay and other species in the Lauraceae in the southeastern United States. References: (1) S. W Fraedrich et al. Plant Dis. 92:215, 2008. (2) T. C. Harrington et al. Mycotaxon 104:399, 2008. (3) M. A. Innis et al. PCR Protocols, A Guide to Methods and Applications. Academic Press. San Diego, CA, 1990. (4) A. E. Mayfield, III et al. Plant Dis. 92:976, 2008.

A Fungal Symbiont of the Redbay Ambrosia Beetle Causes a Lethal Wilt in Redbay and Other Lauraceae in the Southeastern United States.

Extensive mortality of redbay has been observed in the coastal plain counties of Georgia and southeastern South Carolina since 2003 and northeastern Florida since 2005. We show that the redbay mortality is due to a vascular wilt disease caused by an undescribed Raffaelea sp. that is a fungal symbiont of Xyleborus glabratus, an exotic ambrosia beetle. Trees affected by the disease exhibit wilt symptoms that include a black discoloration of the sapwood. Redbay trees and containerized seedlings died within 5 to 12 weeks after inoculation with the Raffaelea sp. When redbay seedlings were challenged with X. glabratus, the beetles tunneled into 96% of the plants, 70% died, and the Raffaelea sp. was recovered from 91%. X. glabratus and the Raffaelea sp. have also been associated with mortality of sassafras, and the Raffaelea sp. has been isolated from wilted pondberry and pondspice. Additional inoculation studies have shown that the Raffaelea sp. is pathogenic to sassafras, spicebush, and avocado, but not to red maple. Female adults of X. glabratus have paired mycangia near the mandibles, and the Raffaelea sp. is routinely isolated from the heads of beetles. The fungus is apparently introduced into healthy redbay during beetle attacks on stems and branches. The wilt currently affecting redbay and sassafras represents a major threat to other members of the Lauraceae indigenous to the Americas, including avocado in commercial production.

First Report of Laurel Wilt Disease Caused by a Raffaelea sp. on Avocado in Florida.

Laurel wilt is a vascular disease of redbay (Persea borbonia (L.) Spreng.) and other plants in the family Lauraceae in the southeastern United States. It is caused by a fungus (Raffaelea sp.) that is vectored by a non-native insect of Asian origin, the redbay ambrosia beetle (Xyleborus glabratus Eichhoff) (1). Since the initial detection of the redbay ambrosia beetle near Savannah, GA in 2002, laurel wilt has caused widespread mortality of redbay in Georgia, South Carolina, and Florida (1). In September 2007, an avocado (Persea americana Mill.) tree planted approximately 10 years earlier in a residential neighborhood in Jacksonville, FL was discovered to be infected with laurel wilt. The crown was in various stages of decline, including upper branches that were dead and leafless, those with wilted and drooping foliage, and those with healthy foliage. Removal of bark from wilted branch sections revealed black-to-brown streaks of discoloration in the sapwood and a few ambrosia beetle holes from which the discoloration extended into the adjacent wood. A Raffaelea sp. was isolated from discolored wood samples by surface sterilizing wood chips by submersion in a 5% sodium hypochlorite solution for 30 s and plating them on cycloheximide streptomycin malt agar (2). Small subunit (18S) sequences from the rDNA were amplified by PCR and sequenced with primers NS1 and NS4 (3). BLASTn searches revealed homology to Raffaelea sp. C2203 (GenBank Accession No. EU123076, 100% similarity, e-value of 0.0, and a total score of 1,886), which is known to be the causal agent of laurel wilt (1). The small-subunit rDNA sequence for this isolate has been deposited into GenBank and has been assigned accession No. EU257806. Pathogenicity of the laurel wilt pathogen on Persea spp. in growth chamber trials has been previously demonstrated (1). Laurel wilt is of concern to the commercial avocado industry and is a potential threat to the Lauraceae elsewhere in the Americas. References: (1) S. W. Fraedrich et al. Plant Dis. 92:215, 2008. (2) T. C. Harrington. Mycologia 73:1123, 1981. (3) T. J. White et al. Page 315 in: PCR Protocols, a Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, San Diego, CA, 1990.