Effects of urban land-use on largescale stonerollers in the Mobile River Basin, Birmingham, AL.

During the spring and fall of 2001 and the spring of 2002 a study was conducted to evaluate the health of the largescale stoneroller (Campostoma oligolepis) populations in streams along an urban land-use gradient. Sites were selected from a pool of naturally similar sub-basins (eco-region, basin size, and geology) of the Mobile River basin (MRB), using an index of urban intensity derived from infrastructure, socioeconomic, and land-use data. This urban land-use gradient (ULUG) is a multimetric indicator of urban intensity, ranging from 0 (background) to 100 (intense urbanization). Campostoma sp. have been used previously as indicators of stream health and are common species found in all sites within the MRB. Endpoints used to determine the effects of urban land-use on the largescale stoneroller included total glutathione, histology, hepatic apoptosis, condition factor and external lesions. Liver glutathione levels were positively associated with increasing urban land-use (r(2) = 0.94). Histopathological examination determined that some abnormalities and lesions were correlated with the ULUG and generally increased in prevalence or severity with increasing urbanization. Liver macrophage aggregates were positively correlated to the ULUG. The occurrence of nucleosomal ladders (indicating apoptotic cell death) did not correspond with urban intensity in a linear fashion. Apoptosis, as well as prevalence and severity of a myxozoan parasite, appeared to have a hormetic dose-response relationship. The majority of the biomarkers suggested fish health was compromised in areas where the ULUG ≥ 36.

First Report of Stemphylium botryosum on Spinach in Texas.

During the 2009 to 2010 growing season, symptoms of an unknown leaf spot were observed on spinach (Spinacia oleracea L.) in production fields in southwest Texas. Approximately 500 ha were affected, especially cvs. Rakaia and Viceroy. Disease incidence was 30 and 2% for Rakaia and Viceroy, respectively. Diseased plants exhibited small (1 to 3 mm in diameter), tan, necrotic lesions with a circular to oval shape and were void of any signs of a pathogen. Symptomatic leaves were surface sterilized in 1.5% NaOCl for 1 min, rinsed with sterile water, and air dried. Leaf sections (~1 cm2) were cut and placed on acidified potato dextrose agar (APDA), or APDA supplemented with streptomycin (SAPDA). Fungal mycelia growing from the edges of infected leaf sections were transferred to PDA and incubated at 25°C with a 12-h/12-h light/dark cycle. After 14 days of incubation, dark brown mycelia giving rise to unbranched conidiophores bearing brown, deeply septate, ovoid conidia were observed. Conidia measured 16.8 to 27.3 × 13.1 to 19.6 µm. On the basis of these morphological characteristics, the fungus was identified as Stemphylium botryosum (3). Cultures were transferred to clarified V8 juice agar to obtain inoculum for pathogenicity tests. Eight-week-old plants (n = 20) of spinach cvs. Hybrid 310, Wintergreen, Ashley, and Rakaia were sprayed until runoff with a suspension containing 0.001% Tween 80 and 1 × 104 conidia/ml. Noninoculated plants served as a control treatment. Plants were placed in a growth chamber and incubated in the dark at 25°C and 95% relative humidity. Following 36 h of incubation, plants were transferred to a plastic enclosure and maintained at 23 ± 4°C. After 7 to 10 days, tan, oval-shaped lesions were observed on all inoculated spinach plants. All control plants, with the exception of Rakaia, failed to develop symptoms. Isolates of S. botryosum were recovered on SAPDA from symptomatic leaves, confirming Koch's postulates. Previous reports have shown that S. botryosum can be transmitted from infected seed (1), thus, additional plants of each cultivar (n = 36) were grown in the greenhouse to determine the potential for seedborne contamination. After 8 weeks, leaf spot symptoms identical to those observed on the original plants developed on 75% of the Rakaia plants, while symptom development on the other cultivars was negligible. Isolates of S. botryosum were only recovered from symptomatic Rakaia leaves. Similar field observations were made during the 2001 to 2002 growing season; however, attempts to isolate S. botryosum in that season were unsuccessful. Recent outbreaks of Stemphylium leaf spot have been reported in Arizona (4), California (3), Delaware and Maryland (2), and Washington (1). To our knowledge, this is the first report of S. botryosum on spinach in Texas. While the origin of inoculum causing the disease in Texas is unknown, S. botryosum may have been seedborne (2). The implementation within the past few years of very high density plantings of spinach (1.9 to 3.7 million seeds/ha) may lead to an increase in incidence and severity of this disease in Texas. References: (1) L. J. du Toit and M. L. Derie. Plant Dis. 85:920, 2001. (2) K. L. Everts and D. K. Armentrout. Plant Dis. 85:1209, 2001. (3) S. T. Koike et al. Plant Dis. 85:126, 2001. (4) S. T. Koike et al. Plant Dis. 89:1359, 2005.

Acute and chronic toxicity of fluoxetine (selective serotonin reuptake inhibitor) in western mosquitofish.

Fluoxetine is a biologically active pharmaceutical chemical that has been detected at parts-per-trillion levels in surface waters in North America and Europe. This has generated concern because negative effects in aquatic organisms are possible. Known as a selective serotonin reuptake inhibitor, fluoxetine (e.g., Prozac; Elli Lilly) is neurologically active and widely prescribed for clinical depression in humans. In the present investigation, acute and chronic toxicities of fluoxetine were evaluated in an environmentally relevant species, western mosquitofish Gambusia affinis. Acute toxicity (5 to 5340 ppb fluoxetine) was assessed in neonates (age 24 to 48 hours) exposed in glass aquaria for 7 days; chronic toxicity (0.05 to 5 ppb fluoxetine) was examined in fish exposed from age neonate to age 91 days; and effects of chronic exposure (100 days) on sexual maturation was investigated in mesocosm tanks (100 L) in fish exposed (7 to 71 ppb) from age 59 to 159 days. Acute toxicity of fluoxetine in neonate western mosquitofish was observed to have a 7-day median lethal concentration of 546 ppb. Chronic exposure did not affect survival, growth, or sex ratio; however, increased lethargy in fish exposed to > or =0.5 ppb fluoxetine was observed. In fish exposed from age 59 to 159 days (juvenile to adult life stages), delayed development of external adult sexual morphology was observed at 71 ppb fluoxetine, which consisted of delayed onset of the presence of the black spot in the posterior abdomen in female fish and delayed formation of the elongated anal fin (gonopodium) in male fish. The present study demonstrated that chronic exposure of western mosquitofish to fluoxetine can affect sexual development; however, it does so only at concentrations 3 to 4 orders of magnitude higher than those previously found in the environment.

Tomato yellow leaf curl virus in Tomato in Texas, Originating from Transplant Facilities.

Tomato yellow leaf curl virus (TYLCV), a monopartite virus in the genus Begomovirus (family, Geminiviridae) from the Middle East, is one of the most damaging whitefly-transmitted viruses of tomato (Lycopersicon esculentum) worldwide. TYLCV was first identified in the United States in 1997 in Florida (4), and most recently, in the Pacific Coast states of Mexico where fresh market tomatoes are grown for the U.S. market (1). During September 2006, tomatoes grown from transplants in Waller County, TX exhibited shortened internodes, stunting and puckering of leaflets, green vein banding, and diffuse chlorosis. The disease incidence in two fields (4 ha total) was 95% and yield was substantially reduced. Many of the transplants were symptomatic at planting. The transplants originated from two facilities in Hidalgo County, TX. Both facilities had experienced heavy infestations of the whitefly, Bemisia tabaci (Genn.), during transplant production. At the same time, transplants produced in Uvalde and Bexar counties, TX, where whitefly infestations were also prevalent, had similar virus symptoms. Total DNA was extracted from the leaves of symptomatic tomato plants from 10 samples from these four counties and amplified by PCR (2). DNA samples from Waller, Hidalgo, and Uvalde counties were cloned, and a partial fragment of the viral coat protein gene (core Cp) was sequenced. BLAST analysis of the core Cp sequences of each sample confirmed the presence of TYLCV. No other begomovirus was detected, and all attempts to amplify a bipartite begomovirus by PCR using degenerate DNA-B specific primers (3) were unsuccessful. The full-length TYLCV DNA was amplified from three samples using the rolling circle amplification method as described (1), cloned, and the sequences were determined. The three sequences shared 99.6 to 100% nt identity and so only one sequence was deposited in the NCBI GenBank database (Accession No. EF110890) (1). Analysis of the complete genome nucleotide sequence corroborated TYLCV identity predicted by core Cp analysis that was 98.1% identical with TYLCV from Egypt (GenBank Accession No. AY594174) and Spain (GenBank Accession No. AJ489258), 97.6% with TYLCV from Mexico (GenBank Accession No. DQ631892), and 96.5% with TYLCV-Is (GenBank Accession No. X15656). Additionally, a Southern blot with TYLCV as the probe detected replicating (double-stranded) TYLCV DNA in all samples consisting of three plants from Uvalde County and 21 plants from Bexar County. To our knowledge, this is the first report of TYLCV in Texas that occurred in two transplant production areas approximately 400 km apart. Transplants produced in Uvalde and Bexar counties were planted there, while Hidalgo County transplants were shipped outside of the usual range of the whitefly. Hidalgo County has a subtropical climate, which can allow overwintering of TYLCV and the whitefly vector, allowing the establishment and spread of this virus in the future. References: (1) J. K. Brown and A. M. Idris. Plant Dis. 90:1360, 2006. (2) J. K. Brown et al. Arch. Virol. 146:1581, 2001. (3) A. M. Idris and J. K. Brown. Phytopathology 88:648, 1998. (4) J. E. Polston et al. Plant Dis. 83:984, 1999.

First Report of Yucca Phyllody Associated with 16SrI-A Phytoplasmas in Texas.

Buckley's yucca (Yucca constricta Buckl.) is a native flowering perennial plant widely distributed in Texas and northeast Mexico. It is also grown as an ornamental plant in its native range as well as in other dry regions in the United States and Mexico. In 2006, during an extended drought, Buckley's yucca plants sporadically exhibited phyllody and abnormal bud proliferation on the inflorescence in Uvalde County in southwestern Texas. Symptoms resembled those caused by phytoplasmal infection. Samples from four symptomatic and two asymptomatic yucca plants were collected. Total nucleic acid was extracted from abnormal bud tissue. To assess the etiological aspect of the disease nested PCR using phytoplasma specific primer pair P1/16S-SR or P1/P7 followed by R16F2n/R16R2n was employed for the detection of putative phytoplasmas (2). An amplicon of approximately 1.2 kb was amplified from all four symptomatic yucca plants but not from asymptomatic plants. Restriction fragment length polymorphism (RFLP) patterns of 16S rDNA digested singly with AluI, KpnI, HpaII, MseI, HhaI, and RsaI endonucleases indicated that affected yucca plants were infected by a phytoplasma belonging to aster yellows group 16SrI ('Candidatus Phytoplasma asteris'), subgroup 16SrI-A (1). Nucleotide sequence analysis of cloned 16S rDNA (GenBank Accession No. EF190067) confirmed the results on the basis of RFLP analyses. Yucca phyllody has not been reported elsewhere. This disease appears to be newly emerging in Texas with only a few affected plants. To our knowledge, this is the first report of 16SrI-A phytoplasma infecting a Yucca sp. References: (1) I.-M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998. (2) I.-M. Lee et al. Int. J. Syst. Evol. Microbiol. 54:337, 2004.

First Report of Iris yellow spot virus on Onion (Allium cepa) in Texas.

Iris yellow spot virus (IYSV; family Bunyaviridae, genus Tospovirus) has emerged as a potentially devastating and widespread virus of onion. IYSV was first reported in the United States from Idaho in 1993 and has since spread to many of the onion-producing areas (1). In South America, the most recent reports of the virus on onion were from Peru and Chile (2,4). In 2005, onion plants in Uvalde County, Texas exhibited necrotic lesions on leaves typical of IYSV and disease incidence approached 100% in some fields with yield loss and quality problems. Five of six plants tested were positive for IYSV with double antibody sandwich-enzyme linked immunosorbent assay (DAS-ELISA; Agdia Inc., Elkhart, IN). In 2006, similar lesions were observed on onion plants in Uvalde County and approximately 400 km south in Hidalgo and Cameron counties. Infection points generally started as a single plant near the edge of fields and spread to plants in a 3- to 4-m area after 1 to 2 weeks. Early-season disease incidence was low in onions grown for bulbs and transplants, <10% in 2006. Disease incidence increased in some fields until the crop was harvested. Leaves of symptomatic plants were tested for IYSV and Tomato spotted wilt virus (TSWV) using DAS-ELISA, and 18 of 23 samples from the Hidalgo County area and 12 of 21 samples from the Uvalde County area were positive for IYSV. All samples tested for TSWV from these counties were negative. Virus infection in some ELISA-positive plants was verified by reverse transcription-polymerase chain reaction (RT-PCR) using primers derived from the small RNA of IYSV. The primers flanked the IYSV nucleocapsid (N) gene (5'-TAA AAC AAA CAT TCA AAC AA-3' and 5'-CTC TTA AAC ACA TTT AAC AAG CAC-3' (3). RT-PCR gave a PCR product of expected size (approximately 1.2 kb). The DNA amplicon was cloned and sequenced (GenBank Accession No. DQ658242). Nucleotide sequence analysis confirmed the identity of the amplicon as that of IYSV N gene and sequence comparisons with known IYSV N gene sequences showed 95 to 98% sequence identity. The primary vector of IYSV, onion thrips (Thrips tabaci), is a widespread and destructive pest of onion in south Texas. The year-to-year incidence of IYSV and the severity of the disease will probably depend on the onion thrips population levels. Bulb yield reduction could be severe during years with high thrips populations. More research is needed to determine the impact of IYSV on bulb yield in Texas, the relationship between IYSV incidence and T. tabaci population levels, and oversummering hosts. To our knowledge, this is the first known report of IYSV in Texas. References: (1) D. H. Gent et al. Plant Dis. 88:446, 2004, (2) S. W. Mullis et al. Plant Dis. 90:377, 2006, (3) H. Pappu et al. Arch. Virol. 151:1015, 2006. (4) M. Rosales et al. Plant Dis. 89:1245, 2005.

First Report of Ramulispora sorghicola in the United States Causing Oval Leaf Spot on Johnsongrass and Sorghum in Texas.

Oval leaf spot (OLS) caused by Ramulispora sorghicola Harris was observed on grain sorghum, Sorghum bicolor (L.) Moench, and johnsongrass, S. halepense (L.) Pers., near Beeville, TX during August 2002. Symptoms were first observed on several sorghum lines and hybrids in a field nursery including a bulk planting of the line ATx623. Highest incidence of OLS occurred in rows adjacent to johnsongrass with symptoms of OLS. Average lesion size (mm) was 1.3 × 2.8 with a range from 0.5 to 2.5 × 1.5 to 5.0. Lesions had a straw-colored sunken center and on red- and purple-pigmented sorghums, lesion borders were highly pigmented. Cone-shaped conidial masses and superficial sclerotia (subglobose, black, 80 to 190 µm in diameter × 50 to 70 µm high, with spiny setae) were sometimes present or readily produced on lesions within 48 to 72 h after placement in humidity chambers. Conidia were branched, filiform, tapered, and 1.1 to 2.4 × 20 to 75 µm. The pathogen, R. sorghicola, was isolated from conidia and sclerotia. A water suspension of culturally derived conidia of R. sorghicola (3 × 104 conidia per ml) was spray inoculated (5:30 p.m., October 11, 2002) onto four or more upper leaves per plant of six grain sorghum plants (ATx623) and approximately nine johnsongrass plants (three tillers each of three plants) at a Corpus Christi field location where OLS was absent. Three grain sorghum and one johnsongrass plant were sprayed with a water control. Cloudy, wet, and cool conditions after inoculation and increasingly cooler nights probably delayed symptom expression until 3 to 4 weeks after inoculation. Typical lesions were observed simultaneously on both hosts with symptoms restricted to inoculated plants. Lesions from both hosts were placed onto water agar at 25°C for 24 h, and the pathogen was reisolated from field-produced conidia of rehydrated conidial masses. Through 2004, OLS was observed on sorghum hosts in 29 counties from central Texas to the Lower Rio Grande Valley. During the growing season, OLS was predominantly absent in grain and forage sorghum fields and absent or often difficult to detect in johnsongrass. In all 3 years, OLS was most common after the normal growing season from August through December with occurrence primarily on johnsongrass but also on late-planted and feral S. bicolor hosts, especially when proximal to symptomatic johnsongrass. Presence and incidence of OLS was highly variable between and within stands of johnsongrass with incidence ranging from a few to most plants. Incidence in forage or grain sorghum fields was highest at field borders adjacent to johnsongrass with OLS. Disease severity was low except on johnsongrass at a few locations. The pathogen appears to pose low economic risk to any sorghum host in Texas at any time of the year although highly susceptible lines and hybrids should be identified and possibly avoided. The previous most proximal report of R. sorghicola in the Western Hemisphere was in Honduras (1). The widespread distribution of OLS across southern Texas and its pattern of occurrence in johnsongrass suggest that the pathogen may have been unobserved in Texas for several years. Presence of OLS near the Rio Grande indicates probable occurrence in johnsongrass at least in some areas along this river in northeastern Mexico. Reference: (1) G. C. Wall et al. Trop. Pest Manag. 35:57, 1989.

First Report of Cylindrocladium Black Rot Caused by Cylindrocladium parasiticum on Peanut in Texas.

During August 2004, Cylindrocladium parasiticum Crous, M.J. Wingf., & Alfenas (teleomorph Calonectria ilicicola Boedijin & Reitsma) was isolated from peanut (Arachis hypogaea L.) cv. NC 7 taken from an irrigated field in Terry County, TX. On 24 September, the mean length of patches with symptoms of dead plants and confirmed for the presence of C. parasiticum was 40 cm (averaged over 52 patches). Pods, pegs, roots, and lower stems of affected plants had decayed black lesions. No perithecia were observed. Roots were plated on water agar, and mycelia growth was transferred to potato dextrose agar (PDA). Conidia and microsclerotia typical of C. parasiticum had formed at 14 days. After 21 days on PDA, the following mean measurements were recorded: macroconidia 57 × 7 µm (usually two septations); stipe 115 × 4 µm; and sphaeropedunculate vesicle 15 × 10 µm. These are within reported ranges for C. parasiticum (1). Four-week-old cultures were comminuted in tap water and added (mycelium and microsclerotia) to potting medium (noninfested nonsterile peanut field soil/peat moss potting mix, 1:1). Peanut-specific Bradyrhizobium sp. was applied before planting. Three 3-day-old peanut cv. Tamrun 88 seedlings were transplanted in infested soil (four replicate pots, five isolates). Root, hypocotyl, pod, peg, and lower stem lesions were evident after 8 (Trial 1, one isolate) and 16 weeks (Trial 2, four isolates). Virulence varied among isolates (3). C. parasiticum was reisolated on acidified PDA after both trials. In Trial 3, six plant species (three replicate pots, five plants per pot) were challenged with one isolate by drenching a tap water suspension of comminuted microsclerotia and mycelium from 4-week-old PDA cultures into potting medium and transplanting peanut seedlings or planting seeds for other hosts. Root rot severity (0 to 5 index) (3) and reisolation percentages were cv. Tamrun 88 peanut 2.0, 100%; cv. Tamrun 96 peanut 2.0, 92%; cv. Hutcheson soybean (Glycine max (L.) Merr.) 3.3, 64%; cv. Blue Lake 274 green bean (Phaseolus vulgaris L.) 2.7, 40%; cv. California Blackeye 8046 southern pea (Vigna unguiculata (L.) Walp. 2.0, 70%; and cv. Jubilee Hybrid sweet corn (Zea mays L.) 0.0, 100%. Noninoculated peanut controls in all trials had no black decayed root lesions and attempts to isolate the fungus were unsuccessful. Peanut seeds planted in the production field were obtained from the southeastern United States. This field had been scheduled for seed production until C. parasiticum was identified. Temperatures and rainfall amounts during the 2004 growing season were less and greater than long-term averages, respectively. No other infested peanut fields have been confirmed or suspected in the region or state. There is continued risk of contamination from shipments of infected seeds from infested areas (2). This pathogen may pose a serious threat to >100,000 ha of peanut production in western Texas and eastern New Mexico. References: (1) P. W. Crous. Taxonomy and Pathology of Cylindrocladium (Calonectria) and Allied Genera. The American Phytopathological Society, St. Paul, MN, 2002. (2) B. L. Randall-Schadel et al. Plant Dis. 85:362, 2001. (3) R. C. Rowe and M. K. Beute. Phytopathology 65:422, 1975.

Effects of chronic cocaine on monoamine levels in discrete brain structures of lactating rat dams.

Chronic gestational cocaine administration has been correlated with high levels of postpartum maternal aggression towards intruders and altered levels of oxytocin in the amygdala. Cocaine may alter both oxytocin and maternal aggression either directly or indirectly through changes in monoamine levels in relevant brain regions. In this study, pregnant female rats were randomly assigned to one of four groups; three cocaine dose groups (7.5, 15 or 30 mg/kg), or a saline-treated group (0.9% normal saline) and given subcutaneous injections twice daily (total volume 2 ml/kg) throughout gestation. Behavioral responses to an inanimate object placed in the homecage were assessed on Postpartum Day (PPD) 6. Immediately following testing, animals were sacrificed and four brain regions implicated in maternal/aggressive behavior (medial preoptic area [MPOA], ventral tegmental area [VTA], hippocampus, and amygdala) were removed for monoamine level analyses using high-performance liquid chromatography. Dams given 30 mg/kg cocaine throughout gestation had significantly higher levels of dopamine (DA) and nonsignificantly elevated serotonin (5-HT) levels relative to saline-treated controls. These dams also exhibited higher frequencies of defensive behavior toward an inanimate object compared to saline-treated controls. Potential mechanisms mediating cocaine-induced increases in responding are proposed.

Ecological implications from a molecular analysis of phytoplasmas involved in an aster yellows epidemic in various crops in Texas.

ABSTRACT In the spring of 2000, an aster yellows (AY) epidemic occurred in carrot crops in the Winter Garden region of southwestern Texas. A survey revealed that vegetable crops, including cabbage, onion, parsley, and dill, and some weeds also were infected by AY phytoplasmas. Nested polymerase chain reaction (PCR) and restriction fragment length polymorphism analysis of PCR-amplified phytoplasma 16S rDNA were employed for the detection and identification of phytoplasmas associated with these crops and weeds. Phytoplasmas belonging to two subgroups, 16SrI-A and 16SrI-B, in the AY group (16SrI), were predominantly detected in infected plants. Carrot, parsley, and dill were infected with both subgroups. Onion and three species of weeds (prickly lettuce, lazy daisy, and false ragweed) were predominantly or exclusively infected by subgroup 16SrI-A phytoplasma strains, while cabbage was infected by subgroup 16SrI-B phytoplasmas. Both types of phytoplasmas were detected in three leafhopper species, Macrosteles fascifrons, Scaphytopius irroratus, and Ceratagallia abrupta, commonly present in this region during the period of the epidemic. Mixed infections were very common in individual carrot, parsley, and dill plants and in individual leafhoppers. Sequence and phylogenetic analyses of 16S rDNA and ribosomal protein (rp) gene sequences indicated that phytoplasma strains within subgroup 16SrI-A or subgroup 16SrI-B, detected in various plant species and putative insect vectors, were highly homogeneous. However, based on rp sequences, two rpI subgroups were identified within the subgroup 16SrI-A strain cluster. The majority of subgroup 16SrI-A phytoplasma strains were classified as rp subgroup rpI-A, but phytoplasma strains detected in one onion sample and two leafhoppers (M. fascifrons and C. abrupta) were different and classified as a new rp subgroup, rpI-N. The degree of genetic homogeneity of the phytoplasmas involved in the epidemic suggested that the phytoplasmas came from the same pool and that all three leafhopper species may have been involved in the epidemic. The different phytoplasma population profiles present in various crops may be attributed to the ecological constraints as a result of the vector-phytoplasma-plant three-way interaction.

Biomarker and bioaccumulation responses of Asian clams exposed to aqueous cadmium.

Measured responses of biochemical or physiological indicators have been suggested to reflect thresholds where pollutants exert their initial effect. Responses in cellulolytic enzyme activity and DNA strand breakage of the Asian clam Corbicula fluminea exposed to cadmium in the laboratory were measured and metal body burdens were determined concurrently. Clams were exposed to aqueous cadmium concentrations of 3, 6, 12, or 25 ppb for 23 and 28 d. Cadmium concentrations in clam tissue were highest in lower cadmium treatments, and body burdens increased with length of exposure in only the 28-d experiment. Cellulolytic enzyme activity decreased with increasing cadmium concentrations for clams in the 28-d experiment. Induced enzyme activities were observed in cadmium treatments for both experiments and are thought to precede declines in activity through the length of exposure. Significant reductions in DNA strand lengths of cadmium exposed clams were observed by wk 3 in the 23-d exposure and by wk 2 in the 28-d exposure. Reduced DNA strand lengths in these cadmium treatments for the 28-d exposure precede significant declines in cellulolytic activity at subsequent sampling events. Combining these data with observations of mortality in higher cadmium treatments suggests that impairment of DNA structural integrity and reduced digestive enzyme activity may indicate metal-induced stress in clams.

First Report of an Aster Yellows Phytoplasma Associated with Cabbage in Southern Texas.

In early spring 2000 carrot crops in southwestern Texas were severely infected by an outbreak of phyllody associated with aster yellows phytoplasma. Cabbage crops that had been planted adjacent to these carrot fields began to display previously unobserved symptoms characteristic of phytoplasma infection. Symptoms included purple discoloration in leaf veins and at the outer edges of leaves on cabbage heads. Proliferation of sprouts also occurred at the base of the stem and between leaf layers of some plants, and sprouts sometimes continued to proliferate on extended stems. About 5% of cabbage plants in the field exhibited these symptoms. Two symptomless and four symptomatic cabbage heads were collected in early April from one cabbage field. Veinal tissues were stripped from each sample and used for total nucleic acid extraction. To obtain specific and sufficient amount of PCR products for analysis, nested PCR was performed by using primer pairs (first with P1/P7 followed by R16F2n/R16R2) (1,2) universal for phytoplasma detection. A specific 16S rDNA fragment (about 1.2 kb) was strongly amplified from the four symptomatic but not from the two asymptomatic samples. The nested PCR products obtained from the four symptomatic samples were then analyzed by restriction fragment length polymorphism (RFLP) using the restriction enzymes MseI, HhaI, and HpaII, and the RFLP patterns were compared to the published patterns of known phytoplasmas (1). The resulting RFLP patterns were identical to those of a phytoplasma belonging to subgroup B of the aster yellows phytoplasma group (16SrI). These RFLP patterns were also evident in putative restriction sites observed in a 1.5 kbp nucleotide sequence of the 16S rDNA. This is the first report of aster yellows phytoplasma associated disease symptoms in cabbage in Texas. The occurrence of cabbage proliferation coincided with the presence of high populations of the insect vector, aster leafhopper. References: (1) I.-M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998. (2) B. Schneider et al. 1995. Molecular and Diagnostic Procedures in Mycoplasmology, Vol. I. Academic Press, San Diego, CA.

First Report of a Member of Aster Yellows Phytoplasma Group and of Clover Proliferation Phytoplasma Group Associated with Onion in Texas.

An unknown disease(s) emerged this spring (2000) in an onion field in southwestern Texas. Infected onion plants exhibited two symptom types, one with shoot proliferation, moderate stunting of plants, and light yellowish discoloration on leaves (A) and the other with only severe stunting of the plants (B). The bulbs of the infected plants collected from both symptom types were smaller than normal. When the aerial shoots were trimmed, the infected (but not asymptomatic or the severely stunted) bulbs produced multiple slender sprouts after storage in room temperature for about a month. These symptoms are characteristic of yellows diseases caused by phytoplasmas. Ten symptomatic (six with symptom type A and four with symptom type B) and ten symptomless onion plants were collected in early May from an onion field about 1 to 2 weeks prior to blooming. Total nucleic acid was extracted from 0.5 g of shoot tissues from each sample. Nested polymerase chain reaction (PCR) using universal primer pairs (P1/P7 followed by R16F2n/R16R2) previously designed based on 16S and 23S rRNA gene sequence (1,2) was employed for the detection of phytoplasma(s) present in the samples. Specific PCR products (all were about 1.2 kb) were heavily amplified from five samples with symptom type A and one with symptom type B. Three of the symptomatic plants showing symptom type B and five of the symptomless samples were scored as weak positives. Restriction fragment length polymorphism (RFLP) analyses of the PCR products obtained from all five symptomatic samples with symptom type A using restriction enzymes including MseI, HhaI, and HpaII revealed that the associated phytoplasmas detected belonged to aster yellows phytoplasma group (16SrI), subgroup A. RFLP analyses of PCR product from the sample with symptom type B indicated that the associated phytoplasma belonged to clover proliferation group (16SrVI), subgroup A (1). Since symptom type A resembles onion yellows reported elsewhere, we propose to adopt "onion yellows" to refer to the new onion disease occurring in Texas. However, correlation between a member of clover proliferation phytoplasma group and onion plants showing severe stunting could not be firmly established. A phytoplasma belonging to 16SrI-B is associated with onion yellows disease reported in Japan (1). This is the first report that onion yellows occurs in the United States. References: (1) I.-M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998. (2) B. Schneider et al. 1995. Molecular and Diagnostic Procedures in Mycoplasmology, Vol. I, Academic Press, San Diego, CA.

Evaluating the efficiency of toxicity abatement in a constructed wetland with Ceriodaphnia dubia.

Constructed wetlands are becoming increasingly popular as low-cost, high-efficiency means of treating agricultural and municipal wastewaters. Monitoring programs for constructed wetlands usually measure physical and chemical characteristics of wetland treatment, including hydraulic residence time and removal of nutrients (N, P), suspended solids, and biochemical oxygen demand (BOD). However, toxicity abatement is seldom measured as evidence of wetland treatment efficiency. In this study, toxicity tests combined with chemical measurements were employed to measure the efficiency of a constructed wetland in treating swine wastes during fall and winter sampling periods. Although the wetland system operated at three wastewater loading rates, only the high-loading-rate cells were tested because of their year-round flows. Wastewater samples were collected prior to, during, and following wetland treatment to track treatment progress as effluents passed through the wetland cells. Toxicity tests with Ceriodaphnia dubia showed significant toxicity abatement of wastewater as it progressed through the constructed wetland system; however, residual toxicity was still observed in the final wetland effluent. No seasonal differences were observed in toxicity abatement between fall and winter wastewater samples, although nitrate and BOD were removed more efficiently during the fall. Results suggest that, while the constructed wetland system is effective in reducing toxicity in swine wastewater, further pre- or posttreatment or additional dilution is necessary before treated effluents are discharged into surface water.

First Report of the Telial Stage of Gymnosporangium exiguum on Ashe Juniper Adjacent to Hawthorn with Rust in Southwest Texas.

The telial stage of Gymnosporangium exiguum (2) on Ashe juniper (Juniperus ashei) was observed in Bandera County, TX, in April and May 1999 after rain events. Symptomatic plants with multiple lesions were found at low frequencies in dense J. ashei populations. Orange telia formed on scales and bark and on 2- to 3-mm-diameter twigs and became nearly inconspicuous when dry. No hypertrophy was observed. Previously reported telial hosts include J. californica and J. excelsa cv. Stricta in California; J. mexicana and J. scopulorum in Oklahoma; J. virginiana in Texas; and J. deppeana var. pachyphloea in Oklahoma and Texas (1). An aecial stage identified as G. exiguum has been observed for many years on native hawthorns (Crataegus crus-galli, C. greggiana, C. mollis, C. stevensiana, C. tracyi, C. turnerorum, C. viridis var. desertorum, and several natural hybrids) in Bandera, Bexar, Edwards, Gillespie, Kendall, Kimble, Real, and Uvalde counties, Texas. Prominent roestelioid aecia were observed on hawthorn leaves, petioles, fruits, peduncles, thorns, and, less often, on twigs. In two experimental plantings of hawthorns in Bandera County, rust severity was rated as low to moderate in six populations of C. greggiana and hybrids; moderate in two populations each of C. stevensiana and C. tracyi; moderate to severe in three populations of C. crus-galli and hybrids; and severe in one population each of C. mollis and C. viridis var. desertorum. G. exiguum was previously reported on a Crataegus sp. in Texas and on Heteromeles arbutifolia in California (1). Flowers, fruits, plant forms, and drought tolerances are characteristics of some endemic hawthorns that provide landscape and wildlife advantages. G. exiguum causing rust disease may limit the ornamental potential of highly susceptible hawthorn species in southwest Texas. Inconspicuous infections on susceptible ornamental Juniperus spp. also could have phytosanitary implications. Voucher specimens (aecia and telia) are on deposit in the Mycological Herbarium, Department of Plant Pathology, North Carolina State University, Raleigh. References: (1) D. F. Farr et al. 1989. Fungi on Plants and Plant Products in the United States. The American Phytopathological Society, St. Paul, MN. (2) F. D. Kern. 1973. A Revised Taxonomic Account of Gymnosporangium. The Pennsylvania State University Press, University Park.

First Report of Natural Infection of Peanut (Groundnut) by Impatiens Necrotic Spot Tospovirus (Family Bunyaviridae).

Impatiens necrotic spot tospovirus (INSV) of the family Bunyaviridae is an important viral pathogen of ornamentals and a major constraint in the greenhouse industry (2). Evidence of natural infection of peanut (groundnut, Arachis hypogaea L.) by INSV was found in samples collected from three sites in Frio County, TX, and one site each in Mitchell and Tift counties, GA, during October 1998. Roots from several plants were tested by enzyme-linked immunosorbent assay for tomato spotted wilt tospovirus (TSWV) and INSV. Symptoms on individual mature plants positive for INSV were the same as those associated with late-season TSWV infections: plants appeared yellow and wilted, internal taproot and crown were necrotic, and plant death resulted (1). At one Texas site, three of five composite samples were positive only for INSV. One composite sample at a second Texas site was positive for both TSWV and INSV. Double infections were found in three of four TSWV-positive samples at a third Texas site. In Mitchell County, GA, three of four samples tested were positive only for TSWV, and one was positive for both TSWV and INSV. In Tift County, GA, 11 of 23 samples tested were positive only for INSV, whereas 4 were positive only for TSWV. Double infections were found in 5 of 23 samples. The presence of INSV in the sample from Mitchell County was verified by immunocapture-polymerase chain reaction (PCR) (4). The apparently low titer of INSV in the doubly infected plant necessitated two cycles of PCR for detection of INSV sequences. A primer pair that can amplify most tospoviruses was used for the first PCR cycle (3). Using the PCR product obtained, a second PCR cycle was performed with one tospovirus-specific and one INSV-specific primer. This approach resulted in a product of the expected size (≈298 bp). The PCR product was cloned in a pGEM-T vector and sequenced. Comparisons indicated the sequence obtained from the infected peanut sample from Georgia was 99% identical to the respective S RNA region from known INSV isolates. Serological and molecular sequence data suggest the peanut samples were infected by INSV. Future surveys and screenings of peanut plants for spotted wilt disease should include a test for INSV. References: (1) A. K. Culbreath et al. Plant Dis. 75:863, 1991. (2) M. Daughtrey et al. Plant Dis. 81:1220, 1997. (3) R. Dewey et al. Virus Genes 13:255, 1996. (4). R. K. Jain et al. Plant Dis. 82:900, 1998.

Biomarker assessment of the effects of coal strip-mine contamination on channel catfish.

A suite of biomarkers was used to evaluate acute (1-day) to semichronic (3-month) heavy metal-induced toxicity in channel catfish, lctalurus punctatus, caged at an abandoned strip mine and a noncontaminated reference site. Assays performed include indicators of metabolic, hematological, osmoregulatory, and genotoxic stress. Two cage designs were used to evaluate the importance of exposure routes: one excluding contact with the sediments and the other allowing contact with water and sediments. Significant DNA strand breakage was observed in catfish exposed to both exposure regimens, but evidence of DNA repair was observed only in water-exposed catfish. Transient increases in hemoglobin, delta-aminolevulinic acid dehydratase, and hematocrit levels were observed at 1 month's exposure for both exposure regimens, followed by a return to control levels for the duration of the study. Environmental conditions (i.e., weather-related changes in water quality) may have contributed to the variable plasma chloride and glucose levels observed in all catfish exposed to strip-mine wastes. The transient changes in biomarkers followed by a return to reference values represent an initial stress and an acclimation to normal levels.

Natural Infection of Citronmelon with Acidovorax avenae subsp. citrulli.

Citronmelon fruits (Citrullus lanatus var. citroides (Bailey) Mansf.) with lesions were collected from a cowpea field in Frio County, TX, in July 1997. The lesions were circular, necrotic, or water-soaked, approximately 3 mm in diameter, and did not extend into the flesh of the fruit. Nonfluorescent, gram-negative bacteria were consistently isolated from lesions. Six representative strains were identified as Acidovorax avenae subsp. citrulli (Aac), using Biolog GN MicroPlates and the MicroLog data base release 3.50 (0.533 to 0.813 similarity). Aac causes leaf and fruit lesions (bacterial fruit blotch, BFB) on watermelon (C. lanatus (Thunb.) Matsum. & Nakai). Strains were tested for pathogenicity on watermelon seedlings (cv. Royal Sweet) by daubing bacterial suspensions (approximately 108 CFU/ml) onto cotyledons of 1-week-old seedlings. Water soaking of cotyledons, followed by necrosis and seedling death, occurred within 5 days. These symptoms were indistinguishable from those caused by watermelon strains of Aac. Bacteria were reisolated from symptomatic seedlings. The source of the infection is not known. Watermelons had been grown in this field in 1996, but no BFB symptoms were observed. Citron fruit infected with Aac were found in nearby watermelon fields where BFB was present; the closest field was 50 m from the cowpea field. These observations suggest that citronmelon, a common weed in south Texas, has the potential to perpetuate Aac. This is the first documentation of a naturally occurring infection of citronmelon with Aac.

Investigations into using the nematode Caenorhabditis elegans for municipal and industrial wastewater toxicity testing.

This investigative study assesses the ease and usefulness of the nematode Caenorhabditis elegans for identifying contributors to effluent toxicity within an industrial and municipal wastewater treatment plant (WWTP) system. Several different types of industries, including fiberglass manufacturing, paper packaging, and yarn dyeing, discharge effluent into the municipal wastewater treatment plant, which in turn discharges into a local creek. A major objective of this study was to identify primary sources of toxicity throughout the system with a nematode toxicity test. Twenty-four-hour composite water samples were taken periodically over a ten-month period at five strategic points within the system: (1) at the point of discharge at each of the three industries, (2) at the combined industrial influent of the wastewater treatment plant, (3) at the effluent of the WWTP, (4) upstream of the WWTP discharge, and (5) downstream of the WWTP discharge. Samples were analyzed for basic water chemistry, and each sample was tested for whole effluent toxicity using a 72-h nematode test with mortality as the end point. Results suggest that interactions between the wastewaters of certain industries may increase the overall nematode toxicity in the wastewater treatment facility's composite influent and effluent. Nematode mortality trends indicate relatively high toxicity levels in wastewater entering the WWTP from contributing industries. High WWTP influent toxicity may potentially be due to varying flow rate ratios of industrial discharges, release of varying toxic constituents in wastewaters, and toxic interactions between chemical constituents of industrial wastewaters. The evaluation of toxicity within the treatment system may pinpoint locations where pollution prevention strategies may be implemented to reduce overall toxicity at the point of discharge.

The effect of space flight on monoclonal antibody synthesis in a hybridoma mouse cell line.

The hybridoma cell line, 3G10G5, producing a monoclonal antibody to the major capsid protein VP1 from the avian polyomavirus budgerigar fledgling disease virus, was produced from a Balb/C mouse. This cell line was used to test the effects of microgravity on cellular processes, specifically protein synthesis. A time course study utilizing incorporation of [35S]methionine into newly synthesized monoclonal antibody was performed on STS-77. After 5.5 days, it was observed that cell counts for the samples exposed to microgravity were lower than those of ground-based samples. However, radiolabel incorporation of the synthesized monoclonal antibody was similar in both orbiter and ground control samples. Overall, microgravity does not seem to have an effect on this cell line's ability to synthesize IgG protein.