Buzz off! An evaluation of ultrasonic acoustic vibration for the disruption of marine micro-organisms on sensor-housing materials.

Biofouling is a process of ecological succession which begins with the attachment and colonization of micro-organisms to a submerged surface. For marine sensors and their housings, biofouling can be one of the principle limitations to long-term deployment and reliability. Conventional antibiofouling strategies using biocides can be hazardous to the environment, and therefore alternative chemical-free methods are preferred. In this study, custom-made testing assemblies were used to evaluate ultrasonic vibration as an antibiofouling process for marine sensor-housing materials over a 28-day time course. Microbial biofouling was measured based on (i) surface coverage, using fluorescence microscopy and (ii) bacterial 16S rDNA gene copies, using Quantitative polymerase chain reaction (PCR). Ultrasonic vibrations (20 KHz, 200 ms pulses at 2-s intervals; total power 16·08 W) significantly reduced the surface coverage on two plastics, poly(methyl methacrylate) and polyvinyl chloride (PVC) for up to 28 days. Bacterial gene copy number was similarly reduced, but the results were only statistically significant for PVC, which displayed the greatest overall resistance to biofouling, regardless of whether ultrasonic vibration was applied. Copper sheet, which has intrinsic biocidal properties was resistant to biofouling during the early stages of the experiment, but inhibited measurements made by PCR and generated inconsistent results later on. SIGNIFICANCE AND IMPACT OF THE STUDY: In this study, ultrasonic acoustic vibration is presented as a chemical-free, ecologically friendly alternative to conventional methods for the perturbation of microbial attachment to submerged surfaces. The results indicate the potential of an ultrasonic antibiofouling method for the disruption of microbial biofilms on marine sensor housings, which is typically a principle limiting factor in their long-term operation in the oceans. With increasing deployment of scientific apparatus in aquatic environments, including further offshore and for longer duration, the identification and evaluation of novel antifouling strategies that do not employ hazardous chemicals are widely sought.

Stripe rust and leaf rust resistance QTL mapping, epistatic interactions, and co-localization with stem rust resistance loci in spring wheat evaluated over three continents.

KEY MESSAGE: In wheat, advantageous gene-rich or pleiotropic regions for stripe, leaf, and stem rust and epistatic interactions between rust resistance loci should be accounted for in plant breeding strategies. Leaf rust (Puccinia triticina Eriks.) and stripe rust (Puccinia striiformis f. tritici Eriks) contribute to major production losses in many regions worldwide. The objectives of this research were to identify and study epistatic interactions of quantitative trait loci (QTL) for stripe and leaf rust resistance in a doubled haploid (DH) population derived from the cross of Canadian wheat cultivars, AC Cadillac and Carberry. The relationship of leaf and stripe rust resistance QTL that co-located with stem rust resistance QTL previously mapped in this population was also investigated. The Carberry/AC Cadillac population was genotyped with DArT(®) and simple sequence repeat markers. The parents and population were phenotyped for stripe rust severity and infection response in field rust nurseries in Kenya (Njoro), Canada (Swift Current), and New Zealand (Lincoln); and for leaf rust severity and infection response in field nurseries in Canada (Swift Current) and New Zealand (Lincoln). AC Cadillac was a source of stripe rust resistance QTL on chromosomes 2A, 2B, 3A, 3B, 5B, and 7B; and Carberry was a source of resistance on chromosomes 2B, 4B, and 7A. AC Cadillac contributed QTL for resistance to leaf rust on chromosome 2A and Carberry contributed QTL on chromosomes 2B and 4B. Stripe rust resistance QTL co-localized with previously reported stem rust resistance QTL on 2B, 3B, and 7B, while leaf rust resistance QTL co-localized with 4B stem rust resistance QTL. Several epistatic interactions were identified both for stripe and leaf rust resistance QTL. We have identified useful combinations of genetic loci with main and epistatic effects. Multiple disease resistance regions identified on chromosomes 2A, 2B, 3B, 4B, 5B, and 7B are prime candidates for further investigation and validation of their broad resistance.

The Pontin series of recombinant alien translocations in bread wheat: single translocations integrating combinations of Bdv2, Lr19 and Sr25 disease-resistance genes from Thinopyrum intermedium and Th. ponticum.

Two bread wheat lines each with a translocation on chromosome 7DL from either Thinopyrum intermedium (TC5 and TC14) or Thinopyrum ponticum (T4m), were hybridized in a ph1b mutant background to enhance recombination between the two translocated chromosomal segments. The frequency of recombinants was high in lines derived from the larger and similar-sized translocations (TC5/T4m), but much lower when derived from different-sized translocations (TC14/T4m). Recombinant translocations contained combinations of resistance genes Bdv2, Lr19 and Sr25 conferring resistance to Barley yellow dwarf virus (BYDV), leaf rust and stem rust, respectively. Their genetic composition was identified using bioassays and molecular markers specific for the two progenitor Thinopyrum species. This set of 7DL Th. ponticum/intermedium recombinant translocations was termed the Pontin series. In addition to Thinopyrum markers, the size of the translocation was estimated with the aid of wheat markers mapped on each of the 7DL deletion bins. Bioassays for BYDV, leaf rust and stem rust were performed under greenhouse and field conditions. Once separated from ph1b background, the Pontin recombinant translocations were stable and showed normal inheritance in successive backcrosses. The reported Pontin translocations integrate important resistance genes in a single linkage block which will allow simultaneous selection of disease resistance. Combinations of Bdv2 + Lr19 or Lr19 + Sr25 in both long and short translocations, are available to date. The smaller Pontins, comprising only 20 % of the distal portion of 7DL, will be most attractive to breeders.

Genetic and QTL analyses of seed dormancy and preharvest sprouting resistance in the wheat germplasm CN10955.

The inheritance and genetic linkage analysis for seed dormancy and preharvest sprouting (PHS) resistance were carried out in an F8 recombinant inbred lines (RILs) derived from the cross between "CN19055" (white-grained, PHS-resistant) with locally adapted Australian cultivar "Annuello" (white-grained, PHS-susceptible). Seed dormancy was assessed as germination index (GI7) while assessment for preharvest sprouting resistance was based on whole head assay (sprouting index, SI) and visibly sprouted seeds (VI). Segregation analysis of the F2, F3 data from the glasshouse and the RIL population in 2004 and 2005 field data sets indicated that seed dormancy and PHS resistance in CN19055 is controlled by at least two genes. Heritabilities for GI7 and VI were high and moderate for SI. The most accurate method for assessing PHS resistance was achieved using VI and GI7 while SI exhibited large genotype by environment interaction. Two quantitative trait loci (QTLs) QPhs.dpivic.4A.1 and QPhs.dpivic.4A.2 were identified. On pooled data across four environments, the major QTL, QPhs.dpivic.4A.2, explained 45% of phenotypic variation for GI7, 43% for VI and 20% for SI, respectively. On the other hand, QPhs.dpivic.4A.1 which accounted for 31% of the phenotypic variation in GI7 in 2004 Horsham field trial, was not stable across environments. Physical mapping of two SSR markers, Xgwm937 and Xgwm894 linked to the major QTL for PHS resistance, using Chinese Spring deletions lines for chromosome 4AS and 4AL revealed that the markers were located in the deletion bins 4AL-12 and 4AL-13. The newly identified SSR markers (Xgwm937/Xgwm894) showed strong association with seed dormancy and PHS resistance in a range of wheat lines reputed to possess PHS resistance. The results suggest that Xgwm937/Xgwm894 could be used in marker-assisted selection (MAS) for incorporating preharvest sprouting resistance into elite wheat cultivars susceptible to PHS.

Is evidence for late selection due to automatic or attentional processing of stimulus identities?

This study determined whether evidence for late selection is due to attention processing or to processing by an automatic system that is separate from attention (two systems framework; Eriksen, Webb, & Fournier, 1990). The task was a two-choice discrimination of a target that appeared in one of two sequentially cued locations in an eight-letter visual display. Attention was directed to the first cued location (cue 1), and whether identification processing occurred at a different location before the second cue (cue 2) directed attention there was determined. Cue validity varied across two experiments, and critical trials were those in which the target appeared at cue 2. For these trials, the target was preceded by a letter (either identical, neutral, or incompatible) that changed to the target at various time intervals following cue 2. Automatic identification was assumed if the incompatible letter interfered with response to the target when it appeared only before cue 2 onset and independent of cue validity. The incompatible letter appearing only before cue 2 onset interfered with the target when the target occurred equally often at cue 1 and cue 2, but not when the target occurred at cue 1 70% and at cue 2 30% of the time. This disconfirms the two systems framework and suggests that attention is required for spatial form processing and response competition.

The stereoscopic (cyclopean) motion aftereffect is dependent upon the temporal frequency of adapting motion.

This study investigated whether the stereoscopic (cyclopean) motion aftereffect (induced by adaptation to moving binocular disparity information) is dependent upon the temporal frequency or speed of adapting motion. The stereoscopic stimuli were gratings created from disparity embedded in a dynamic random-dot stereogram. Across different combinations of stereoscopic spatial frequency, temporal frequency and speed of adapting motion, the duration of the aftereffect was dependent upon temporal frequency (maximal aftereffect=1-2 cyc s(-1)). These results support the idea that stereoscopic motion is processed by a cortical mechanism that computes cyclopean motion energy.

Exposure duration affects the perceived direction of cyclopean type II plaids.

This study investigated the effect of exposure duration on the perceived direction of cyclopean Type I and Type II plaids moving in the X/Y plane. The cyclopean plaids were created from grating components defined by binocular disparity embedded in a dynamic random-dot stereogram. The results showed that the cyclopean Type I plaid appeared to move in the intersection-of-constraints (IOC) direction across the range of exposures tested. However, the cyclopean Type II plaids appeared to move in a direction different from the IOC with short exposures but near the IOC with long exposures. This perceived directional shift was also obtained with luminance-defined Type II plaids. A common pattern-motion mechanism that processes cyclopean and luminance motion signals appears responsible for the perceived directional shift of the Type II plaids.

Cross-domain adaptation reveals that a common mechanism computes stereoscopic (cyclopean) and luminance plaid motion.

Across three experiments, this study investigated the visual processing of moving stereoscopic plaid patterns (plaids created with cyclopean components defined by moving binocular disparity embedded in a dynamic random-dot stereogram). Results showed that adaptation to a moving stereoscopic plaid or its components affected the perceived coherence of a luminance test plaid, and vice versa. Cross-domain adaptation suggests that stereoscopic and luminance motion signals feed into a common pattern-motion mechanism, consistent with the idea that stereoscopic motion signals are computed early in the motion processing stream.

Occlusion contributes to temporal processing differences between crossed and uncrossed stereopsis in random-dot displays.

Stereoscopic depth discrimination was investigated in crossed and uncrossed directions using stimuli defined by binocular disparity differences embedded in dynamic random-dot stereograms. Across three experiments, fixation was directed to a point on the display screen (which placed crossed stimuli in front of and uncrossed stimuli behind, the background dots of the stereogram), to a point in front of the display screen (which placed both crossed and uncrossed stimuli in front of the background dots), and to a point behind the display screen (which placed both crossed and uncrossed stimuli behind the background dots). Results showed that depth discrimination was always good when the stimuli appeared in front of the background dots of the stereogram, whereas discrimination was always poor when the stimuli appeared behind the background dots. These results suggest that differences between crossed and uncrossed stereopsis as reported in past research arose, in part, from effects related to occlusion.

The stereoscopic (cyclopean) motion aftereffect is selective for spatial frequency and orientation of disparity modulation.

Across two experiments, this study investigated the spatial frequency tuning and orientation tuning (both in the disparity domain) of the stereoscopic (cyclopean) motion aftereffect. In Experiment 1, observers adapted to a moving stereoscopic grating of a given cyclopean spatial frequency and tested for the motion aftereffect with a static grating of the same or different spatial frequency. Robust motion aftereffects were induced only when the spatial frequency of the adapt and test stimuli was the same. In Experiment 2, observers adapted to a moving stereoscopic grating of a given cyclopean orientation and tested for the motion aftereffect with a static grating of the same or different orientation. Robust motion aftereffects were induced only when the orientation of the adapt and test stimuli was the same. Together, these results suggest that the stereoscopic motion aftereffect is tuned for cyclopean spatial frequency and orientation which, in turn, suggest that the stereoscopic motion aftereffect is mediated by low-level oriented spatial-frequency mechanisms.

HLA-G isoforms produced by placental cytotrophoblasts and found in amniotic fluid are due to unusual glycosylation.

The human placenta expresses HLA-G, a nonclassical (class Ib) MHC molecule that could play a central role in maternal tolerance of the semiallogeneic fetus. In this work, we report the production of a new mAb, 4H84, that specifically reacts with HLA-G in two formats: immunocytochemistry and immunoblotting. Immunolocalization experiments with 4H84 confirmed our previous finding that cytotrophoblasts within the uterine wall are the only cells in tissue sections of placenta that express the HLA-G protein. Additional experiments showed that both amniocytes and cytotrophoblasts in the amnion-chorion express this protein. Since multiple HLA-G transcripts have been described, we used immunoblotting to study the HLA-G isoforms produced by cytotrophoblasts in vitro and by the amnion-chorion in vivo. Cytotrophoblasts, their conditioned medium, and amniotic fluid samples contained heterodisperse immunoreactive bands (Mr 35,000-50,000). N-deglycosylation by peptide-N-glycosidase F digestion resolved these isoforms into two distinct bands. Cell samples contained primarily an Mr 37,000-42,000 protein, most likely encoded by the full-length mRNA. Conditioned medium and amniotic fluid contained a slightly smaller protein, most likely the secreted form lacking the transmembrane and cytoplasmic regions. Removal of polylactosamine chains by endo-beta D-galactosidase digestion significantly reduced the electrophoretic mobility of the immunoreactive bands, suggesting that HLA-G, unlike class Ib molecules studied to date, carries N-acetyllactosamine units. These data show that Mr heterogeneity of HLA-G is due to its novel glycosylation, rather than to the translation of alternatively spliced mRNAs. We postulate that the unusual carbohydrate structures this molecule carries could interact with maternal immune cells and/or stabilize the molecule.

Expression of granulocyte-colony stimulating factor and its receptor is regulated during the development of the human placenta.

The development of the placenta is dependent upon the regulated proliferation, invasion and differentiation of trophoblast. Expression of cytokines at the feto-maternal interface suggests that these molecules may participate in placentation. The expression of granulocyte-colony stimulating factor (G-CSF) and G-CSF receptor (G-CSFR) during the development of the human placenta was studied by immunohistochemistry using an anti-G-CSF monoclonal antibody (mAb) and two novel anti-G-CSFR mAbs. G-CSF was present in the stroma of fetal chorionic villi and maternal decidual tissues throughout pregnancy. G-CSFR was detected at high levels in fetal first and third, but not second trimester placental tissues. Staining for G-CSFR was undetectable in maternal decidual tissue from all gestational stages. In first trimester tissues, staining for placental G-CSFR was strongest in differentiated syncytiotrophoblast and invasive, extravillous cytotrophoblast, and weak staining was evident in undifferentiated cytotrophoblast. Immunohistochemical data suggesting temporal regulation of G-CSFR were corroborated by Western blotting and amplification by reverse transcription and PCR of G-CSFR mRNA. These data suggested that expression of G-CSFR in the human placenta is regulated both temporally and spatially, and that placental G-CSF is involved in paracrine regulation, and indicate a role for G-CSF and G-CSFR in trophoblast growth or function during placentation.

Antigenic heterogeneity of human cytotrophoblast and evidence for the transient expression of MHC class I antigens distinct from HLA-G.

Expression of MHC class I antigens on trophoblast populations in first trimester human chorionic villous tissue was assessed by immunohistology. Antibodies used were W6/32 which recognizes a non-polymorphic framework determinant of HLA- A, -B, -C, MHM5 specific for HLA-B, C and 4E and B23.1 which are specific for HLA-B. Syncytiotrophoblast and villous cytotrophoblast were negative with all the anti (HLA class I) antibodies tested. Interstitial trophoblast cells within the maternal decidua were identified with a new antibody, NDOG5, which is specific for extravillous cytotrophoblast. Double labelling showed that they bind W6/32 but not 4E, MHM5 or B23.1; consistent with the expression of the monomorphic HLA-G. In contrast the cytotrophoblast cells of the cell islands and cytotrophoblast shell, which also express the NDOG5 antigen, were positive with W6/32, 4E, MHM5 and B23.1. Cell column cytotrophoblast cells were negative with all four MHC class I antibodies. These results suggest that differentiation of cytotrophoblast from noninvasive to invasive forms is associated with transient expression of class I antigens other than HLA-G on cytotrophoblast shell and cell island cytotrophoblast.

First year of AIDS services delivery under Title I of the Ryan White CARE Act.

This is a review of (a) the emergency assistance for ambulatory HIV medical and support services provided in the first year by eligible metropolitan areas (EMAs) funded under Title I of the Ryan White Comprehensive AIDS Resources Emergency (CARE) Act of 1990, (b) the varied responses and processes by which the 16 urban areas receiving Title I funds in 1991 met legislative mandates, (c) the central nature of planning councils under Title I and their formation and functioning, and (d) issues related to current implementation and future expansion of Title I to additional eligible metropolitan areas. Integral to the review is a brief discussion of the history of AIDS and HIV infection, particularly in cities receiving CARE Act funding, an overview of Title I requirements, and a description of the organizational structures cities are using to implement Title I. Information on Title I EMAs is based on analysis of their 1991 applications, bylaws of their HIV service planning councils, intergovernmental agreements between Title I cities and other political entities, and contracts executed by Title I grantees with providers for the delivery of services. Interviews with personnel in several Title I EMAs, including planning council members and grantee staff members, provided additional information. This is the first descriptive accounting of activities related to the 1991 applications for and uses of Title I funds, and the administrative and service issues related to this process.

Localization of tumour necrosis factor production in cells at the materno/fetal interface in human pregnancy.

Biologically active tumour necrosis factor (TNF) was detected in medium conditioned by incubation with explants of human pregnancy decidua or fetal chorionic villous tissue, taken in the first trimester and at term. Addition of endotoxin increased TNF release in most cases. ELISA assays gave similar results for TNF-alpha and also demonstrated low levels of TNF-beta. Using cell populations purified by flow cytometry, secretion of biologically active TNF was shown to be localized to the macrophages. Cytotrophoblast purified from term amniochorion produced no TNF. Both decidual and chorionic villous tissue at term contained mRNA for TNF-alpha and TNF-beta. TNF-alpha mRNA was confined to decidual macrophages in first trimester tissue, and was not present in chorionic cytotrophoblast. TNF-beta mRNA, in contrast, was detected in both macrophage and non-macrophage populations in term decidua.

Production of granulocyte colony-stimulating factor at the materno-foetal interface in human pregnancy.

A bioassay specific for human granulocyte colony-stimulating factor (G-CSF) was developed and used to measure G-CSF production in human pregnancy tissues. G-CSF was secreted by both foetal chorionic villous and maternal decidual tissues taken in the first trimester and at term. The level of G-CSF production by placental tissue was 6750 (1250-10,000) units of bioactivity per g of tissue in 48 hr in the first trimester and 104 (83-190) U/g at term. Bioactive G-CSF was also secreted by decidual tissue, more in the first trimester than at term. ELISA immunoassays measured 75 (10-820) ng/g/48 hr of G-CSF antigen from first trimester placenta, 15 (10-50) ng/g from first trimester decidua and less than 2 ng/g from term placenta. RNA isolated from decidual and chorionic villous tissue or from cells purified by flow cytometry, contained G-CSF mRNA in both tissues. In decidua, mRNA for G-CSF was confined to the macrophages, and cytotrophoblast from term amniochorion contained no detectable G-CSF mRNA. No G-CSF, measured as bioactivity or as mRNA, was detectable in choriocarcinoma cell lines.

Isolation and purification of human endometrial stromal and glandular cells using immunomagnetic microspheres.

Isolation of pure preparations of the different cell populations of human endometrium is a prerequisite for studies of in-vitro function. Sieving of dispersed endometrial cells, followed by adsorption onto immunomagnetic microspheres coated with antibody to Thy-1 was used to separate glandular and stromal cells. The purity of these cell populations was checked with antibodies to cytokeratin and Thy-1. The stromal cells were 98% pure and 90% viable, gland cells were 82% pure with 76% viability. The purified cells were able to proliferate in vitro as shown by thymidine incorporation.

Purification of human cytotrophoblast from term amniochorion by flow cytometry.

Term cytotrophoblast do not express polymorphic MHC Class I antigens, unlike other fetal and maternal cells in the amniochorion/decidua. This allows cytotrophoblast to be isolated and purified from this tissue, utilizing 4E, a monoclonal antibody specific for HLA-B, which labels only non-trophoblast. We have developed a method using enzymic dispersion and Percoll gradient centrifugation, followed by flow cytometry, that yields, on average, a total of 5 X 10(6) term extravillous cytotrophoblast, 97 per cent pure. The availability of highly purified extravillous cytotrophoblast, for the first time, permits precise investigation of trophoblast function.