bluuebunny/crossref_metadata_2025 huggingface dataset:
ABA depolarizes guard cells in intact plants, through a transient activation of R‐ and S‐type anion channels
Authors: Roelfsema M. Rob G. , Levchenko Victor , Hedrich Rainer
Published: December 2004

During drought, the plant hormone abscisic acid (ABA) induces rapid stomatal closure and in turn reduces transpiration. Stomatal closure is accompanied by large ion fluxes across the plasma membrane, carried by K+ and anion channels. We recorded changes in the activity of these channels induced by ABA, for guard cells of intact Vicia faba plants. Guard cells in their natural environment were impaled with double‐barrelled electrodes, and ABA was applied via the leaf surface. In 45 out of 85 cells tested, ABA triggered a transient depolarization of the plasma membrane. In these cells, the membrane potential partially recovered in the presence of ABA; however, a full recovery of the membrane potentials was only observed after removal of ABA. Repetitive ABA responses could be evoked in single cells, but the magnitude of the response varied from one hormone application to the other. The transient depolarization correlated with the activation of anion channels, which peaked 5 min after introduction of the stimulus. In guard cells with a moderate increase in plasma membrane conductance (ΔG < 5 nS), ABA predominantly activated voltage‐independent (slow (S)‐type) anion channels. During strong responses (ΔG > 5 nS), however, ABA activated voltage‐dependent (rapid (R)‐type) in addition to S‐type anion channels. We conclude that the combined activation of these two channel types leads to the transient depolarization of guard cells. The nature of this ABA response correlates with the transient extrusion of Cl− from guard cells and a rapid but confined reduction in stomatal aperture.
Activation of a COI1‐dependent pathway in Arabidopsis by Pseudomonas syringae type III effectors and coronatine
Authors: He Ping , Chintamanani Satya , Chen Zhongying , Zhu Lihuang , Kunkel Barbara N. , Alfano James R. , Tang Xiaoyan , Zhou Jian‐Min
Published: December 2004

Gram‐negative bacteria use a variety of virulence factors including phytotoxins, exopolysaccharides, effectors secreted by the type III secretion system, and cell‐wall‐degrading enzymes to promote parasitism in plants. However, little is known about how these virulence factors alter plant cellular responses to promote disease. In this study, we show that virulent Pseudomonas syringae strains activate the transcription of an Arabidopsis ethylene response factor (ERF) gene, RAP2.6, in a coronatine insensitive 1 (COI1)‐dependent manner. A highly sensitive RAP2.6 promoter‐firefly luciferase (RAP2.6‐LUC) reporter line was developed to monitor activities of various bacterial virulence genes. Analyses of P. syringae pv. tomato DC3000 mutants indicated that both type III secretion system and the phytotoxin coronatine are required for RAP2.6 induction. We show that at least five individual type III effectors, avirulence B (AvrB), AvrRpt2, AvrPphB, HopPtoK, and AvrPphEPto, contributed to RAP2.6 induction. Gene‐for‐gene recognition was not involved in RAP2.6 induction because plants lacking RPM1 and RPS2 responded normally to AvrB and AvrRpt2 in RAP2.6 expression. Interestingly, the role of coronatine in RAP2.6 induction can be partially substituted by the addition of avrB in DC3000, suggesting that AvrB may mimic coronatine. These results suggest that P. syringae type III effectors and coronatine act by augmenting a COI1‐dependent pathway to promote parasitism.
Functional analyses of the flowering time geneOsMADS50, the putativeSUPPRESSOR OF OVEREXPRESSION OF CO 1/AGAMOUS‐LIKE 20(SOC1/AGL20) ortholog in rice
Authors: Lee Shinyoung , Kim Joonyul , Han Jong‐Jin , Han Min‐Jung , An Gynheung
Published: December 2004

A late‐flowering mutant was isolated from rice T‐DNA‐tagging lines. T‐DNA had been integrated into the K‐box region ofOryza sativa MADS50(OsMADS50), which shares 50.6% amino acid identity with theArabidopsisMADS‐box geneSUPPRESSOR OF OVEREXPRESSION OF CO 1/AGAMOUS‐LIKE 20(SOC1/AGL20). While overexpression ofOsMADS50caused extremely early flowering at the callus stage,OsMADS50RNAi plants exhibited phenotypes of late flowering and an increase in the number of elongated internodes. This confirmed that the phenotypes observed in the knockout (KO) plants are because of the mutation inOsMADS50. RT‐PCR analyses of theOsMADS50KO and ubiquitin (ubi):OsMADS50plants showed thatOsMADS50is an upstream regulator ofOsMADS1,OsMADS14,OsMADS15,OsMADS18, andHd (Heading date)3a, but works either parallel with or downstream ofHd1andO. sativa GIGANTEA(OsGI). These results suggest thatOsMADS50is an important flowering activator that controls various floral regulators in rice.
Expression and function of HD2‐type histone deacetylases in Arabidopsis development
Authors: Zhou Changhe , Labbe Helene , Sridha Sunandini , Wang Li , Tian Lining , Latoszek‐Green Marysia , Yang Zhen , Brown Daniel , Miki Brian , Wu Keqiang
Published: December 2004

HD2 (histone deacetylase) proteins are plant‐specific histone deacetylases (HDACs). The Arabidopsis genome contains four HD2 genes, namely HD2A, HD2B, HD2C, and HD2D. We have previously demonstrated that HD2A, HD2B, and HD2C can repress transcription directly by targeting to promoters in planta. Here, we show that the N‐terminal conserved motif (EFWG) and histidine 25 (H25), a potential catalytic residue, were important for the gene repression activity of HD2A. In situ hybridization indicated that HD2A, HD2B, and HD2C were expressed in ovules, embryos, shoot apical meristems, and primary leaves. Furthermore, all three genes were strongly induced during the process of somatic embryogenesis. HD2D mRNA was only detected in the stems and flowers with young siliques and may have adopted different functions. Using green fluorescent protein (GFP) fusions, we demonstrated that HD2A, HD2B, and HD2C accumulated in the nuclei of Arabidopsis cells. Overexpression of 35S::GFP–HD2A in transgenic Arabidopsis plants generated pleiotropic developmental abnormalities, including abnormal leaves, delayed flowering, and aborted seed development. The data showed that normal pattern of HD2 expression was essential for normal plant development and that HD2A, HD2B, and HD2C may be needed for embryogenesis and embryo development. Reverse transcriptase (RT)‐PCR analysis revealed that a number of genes involved in seed development and maturation were repressed in the 35S::GFP–HD2A plants, supporting a role of HD2A in the regulation of gene expression during seed development.
Expression profiling of phyB mutant demonstrates substantial contribution of other phytochromes to red‐light‐regulated gene expression during seedling de‐etiolation
Authors: Tepperman James M. , Hudson Matthew E. , Khanna Rajnish , Zhu Tong , Chang Sherman H. , Wang Xun , Quail Peter H.
Published: December 2004

Different Arabidopsis phytochrome (phy) family members (phyA through phyE) display differential photosensory and/or physiological functions in regulating growth and developmental responses to light signals. To identify the genes regulated by phyB in response to continuous monochromatic red light (Rc) during the induction of seedling de‐etiolation, we have performed time‐course, microarray‐based expression profiling of wild type (WT) and phyB null mutants. Comparison of the observed expression patterns with those induced by continuous monochromatic far‐red light (FRc; perceived exclusively by phyA) in WT and phyA null‐mutant seedlings suggests early convergence of the FRc and Rc photosensory pathways to control a largely common transcriptional network. phyB mutant seedlings retain a surprisingly high level of responsiveness to Rc for the majority of Rc‐regulated genes on the microarray, indicating that one or more other phys have a major role in regulating their expression. Combined with the robust visible morphogenic phenotype of the phyB mutant in Rc, these data suggest that different members of the phy family act in organ‐specific fashion in regulating seedling de‐etiolation. Specifically, phyB appears to be the dominant, if not exclusive, photoreceptor in regulating a minority population of genes involved in suppression of hypocotyl cell elongation in response to Rc signals. By contrast, this sensory function is apparently shared by one or more other phys in regulating the majority Rc‐responsive gene set involved in other important facets of the de‐etiolation process in the apical region, such as cotyledon cell expansion.
Drought tolerance established by enhanced expression of the CC–NBS–LRR gene, ADR1, requires salicylic acid, EDS1 and ABI1
Authors: Chini Andrea , Grant John J. , Seki Motoaki , Shinozaki Kazuo , Loake Gary J.
Published: December 2004

An activation‐tagged allele of activated disease resistance 1 (ADR1) has previously been shown to convey broad spectrum disease resistance. ADR1 was found to encode a coiled‐coil (CC)–nucleotide‐binding site (NBS)–leucine‐rich repeat (LRR) protein, which possessed domains of homology with serine/threonine protein kinases. Here, we show that either constitutive or conditional enhanced expression of ADR1 conferred significant drought tolerance. This was not a general feature of defence‐related mutants because cir (constitutive induced resistance)1, cir2 and cpr (constitutive expressor of PR genes)1, which constitutively express systemic acquired resistance (SAR), failed to exhibit this phenotype. Cross‐tolerance was not a characteristic of adr1 plants, rather they showed increased sensitivity to thermal and salinity stress. Hence, adr1‐activated signalling may antagonise some stress responses. Northern analysis of abiotic marker genes revealed that dehydration‐responsive element (DRE)B2A but not DREB1A, RD (response to dehydration)29A or RD22 was expressed in adr1 plant lines. Furthermore, DREB2A expression was salicylic acid (SA) dependent but NPR (non‐expressor of PR genes)1 independent. In adr1/ADR1 nahG (naphthalene hydroxylase G), adr1/ADR1 eds (enhanced disease susceptibility)1 and adr1/ADR1 abi1 double mutants, drought tolerance was significantly reduced. Microarray analyses of plants containing a conditional adr1 allele demonstrated that a significant number of the upregulated genes had been previously implicated in responses to dehydration. Therefore, biotic and abiotic signalling pathways may share multiple nodes and their outputs may have significant functional overlap.
Light induces phenylpropanoid metabolism in Arabidopsis roots
Authors: Hemm Matthew R. , Rider Stanley D. , Ogas Joseph , Murry Daryl J. , Chapple Clint
Published: December 2004

Experiments have shown that many phenylpropanoid genes are highly expressed in light‐grown Arabidopsis roots. Studies employing reporter gene constructs have indicated that the expression of these genes is localized not only to the lignifying root vasculature, but also to non‐lignifying tissues, such as the root cortex, suggesting that the proteins encoded by these genes may be involved in aspects of phenylpropanoid metabolism other than lignification. Consistent with this hypothesis, roots of etiolated and soil‐grown plants contain almost no soluble phenylpropanoids, but exposure to light leads to the accumulation of flavonoids, as well as high levels of coniferin and syringin (coniferyl and sinapyl‐4‐O‐glycosides), compounds not previously reported to be accumulated in Arabidopsis. To elucidate the mechanism by which light induces root secondary metabolism, extracts of mutants defective in light perception and light responses were analyzed for phenylpropanoid content. The results of these assays showed that phytochrome (PHY)B and cryptochrome (CRY)2 are the primary photoreceptors involved in light‐dependent phenylpropanoid accumulation, and that the hypocotyl elongated (HY5) transcription factor is also required for this response. The presence of phenylpropanoids in etiolated roots of cop (constitutively photomorphogenic)1, cop9, and det (de‐etiolated)1 mutants indicate that the corresponding wild‐type genes are required to repress root phenylpropanoid biosynthesis in the absence of light. Biochemical analysis of root cell walls and analysis of phenylpropanoid gene expression suggest that coniferin and syringin accumulation may be the result of both increased biosynthesis and decreased conversion of these compounds into other phenylpropanoid end products. Finally, our data suggest that the accumulation of coniferin, syringin, and flavonoids in Arabidopsis roots is a high‐irradiance response (HIR), and suggest that comparative analysis of light‐ and dark‐grown Arabidopsis roots may provide new insights into both phenylpropanoid biosynthesis and light signaling in plants.
An intragenic tandem duplication in a transcriptional regulatory gene for anthocyanin biosynthesis confers pale‐colored flowers and seeds with fine spots in Ipomoea tricolor
Authors: Park Kyeung‐Il , Choi Jeong‐Doo , Hoshino Atsushi , Morita Yasumasa , Iida Shigeru
Published: December 2004

While the wild‐type morning glory (Ipomoea tricolor) displays bright‐blue flowers and dark‐brown seeds, its spontaneous mutant, Blue Star, carrying the mutable ivory seed‐variegated (ivs‐v) allele, exhibits pale‐blue flowers with a few fine blue spots and ivory seeds with tiny dark‐brown spots. The mutable allele is caused by an intragenic tandem duplication of 3.3 kbp within a gene for transcriptional activator containing a basic helix‐loop‐helix (bHLH) DNA‐binding motif. Each of the tandem repeats is flanked by a 3‐bp sequence AAT, indicating that the 3‐bp microhomology is used to generate the tandem duplication. The transcripts in the pale‐blue flower buds of the mutant contain an internal 583‐bp tandem duplication that results in the production of a truncated polypeptide lacking the bHLH domain. The mRNA accumulation of most of the structural genes encoding enzymes for anthocyanin biosynthesis in the flower buds of the mutant was significantly reduced. The transcripts identical to the wild‐type mRNAs for the transcriptional activator were present abundantly in blue spots of the variegated flowers, whereas the transcripts containing the 583‐bp tandem duplication were predominant in the pale‐blue background of the same flowers. The flower and seed variegations studied here are likely to be caused by somatic homologous recombination between an intragenic tandem duplication in the gene encoding a bHLH transcriptional activator for anthocyanin biosynthesis, whereas various flower variegations are reported to be caused by excision of DNA transposons inserted into pigmentation genes.
A compendium of methods useful for characterizing Arabidopsis pollen mutants and gametophytically‐ expressed genes
Authors: Johnson‐Brousseau Sheila A. , McCormick Sheila
Published: December 2004

This article provides detailed protocols for collecting pollen and outlines genetic crosses and phenotypic assays that are useful for characterizing mutants that affect pollen development.
Detection of protein–protein interactions in plants using bimolecular fluorescence complementation
Authors: Bracha‐Drori Keren , Shichrur Keren , Katz Aviva , Oliva Moran , Angelovici Ruthie , Yalovsky Shaul , Ohad Nir
Published: December 2004

Protein function is often mediated via formation of stable or transient complexes. Here we report the determination of protein–protein interactions in plants using bimolecular fluorescence complementation (BiFC). The yellow fluorescent protein (YFP) was split into two non‐overlapping N‐terminal (YN) and C‐terminal (YC) fragments. Each fragment was cloned in‐frame to a gene of interest, enabling expression of fusion proteins. To demonstrate the feasibility of BiFC in plants, two pairs of interacting proteins were utilized: (i) the α and β subunits of the Arabidopsis protein farnesyltransferase (PFT), and (ii) the polycomb proteins, FERTILIZATION‐INDEPENDENT ENDOSPERM (FIE) and MEDEA (MEA). Members of each protein pair were transiently co‐expressed in leaf epidermal cells of Nicotiana benthamiana or Arabidopsis. Reconstitution of a fluorescing YFP chromophore occurred only when the inquest proteins interacted. No fluorescence was detected following co‐expression of free non‐fused YN and YC or non‐interacting protein pairs. Yellow fluorescence was detected in the cytoplasm of cells that expressed PFT α and β subunits, or in nuclei and cytoplasm of cells that expressed FIE and MEA. In vivo measurements of fluorescence spectra emitted from reconstituted YFPs were identical to that of a non‐split YFP, confirming reconstitution of the chromophore. Expression of the inquest proteins was verified by immunoblot analysis using monoclonal antibodies directed against tags within the hybrid proteins. In addition, protein interactions were confirmed by immunoprecipitations. These results demonstrate that plant BiFC is a simple, reliable and relatively fast method for determining protein–protein interactions in plants.
The BOS loci of Arabidopsis are required for resistance to Botrytis cinerea infection
Authors: Veronese Paola , Chen Xi , Bluhm Burton , Salmeron John , Dietrich Robert , Mengiste Tesfaye
Published: December 2004

Three Botrytis‐susceptible mutants bos2, bos3, and bos4 which define independent and novel genetic loci required for Arabidopsis resistance to Botrytis cinerea were isolated. The bos2 mutant is susceptible to B. cinerea but retains wild‐type levels of resistance to other pathogens tested, indicative of a defect in a response pathway more specific to B. cinerea. The bos3 and bos4 mutants also show increased susceptibility to Alternaria brassicicola, another necrotrophic pathogen, suggesting a broader role for these loci in resistance. bos4 shows the broadest range of effects on resistance, being more susceptible to avirulent strain of Pseudomonas syringae pv. tomato. Interestingly, bos3 is more resistant than wild‐type plants to virulent strains of the biotrophic pathogen Peronospora parasitica and the bacterial pathogen P. syringae pv. tomato. The Pathogenesis Related gene 1 (PR‐1), a molecular marker of the salicylic acid (SA)‐dependent resistance pathway, shows a wild‐type pattern of expression in bos2, while in bos3 this gene was expressed at elevated levels, both constitutively and in response to pathogen challenge. In bos4 plants, PR‐1 expression was reduced compared with wild type in response to B. cinerea and SA. In bos3, the mutant most susceptible to B. cinerea and with the highest expression of PR‐1, removal of SA resulted in reduced PR‐1 expression but no change to the B. cinerea response. Expression of the plant defensin gene PDF1‐2 was generally lower in bos mutants compared with wild‐type plants, with a particularly strong reduction in bos3. Production of the phytoalexin camalexin is another well‐characterized plant defense response. The bos2 and bos4 mutants accumulate reduced levels of camalexin whereas bos3 accumulates significantly higher levels of camalexin than wild‐type plants in response to B. cinerea. The BOS2, BOS3, and BOS4 loci may affect camalexin levels and responsiveness to ethylene and jasmonate. The three new mutants appear to mediate disease responses through mechanisms independent of the previously described BOS1 gene. Based on the differences in the phenotypes of the bos mutants, it appears that they affect different points in defense response pathways.
Virus‐induced gene silencing as a tool for functional genomics in a legume species
Authors: Constantin Gabriela D. , Krath Britta N. , MacFarlane Stuart A. , Nicolaisen Mogens , Elisabeth Johansen Ida , Lund Ole S.
Published: December 2004

Virus‐induced gene silencing (VIGS) is an attractive reverse‐genetics tool for studies of gene function. However, efficient VIGS has only been accomplished in a few plant species. In order to extend the application of VIGS, we examined whether a VIGS vector based on Pea early browning virus (PEBV) would produce recognizable phenotypes in Pisum sativum. A plasmid vector of PEBV was modified to allow agro‐inoculation and insertion of heterologous sequences. cDNA fragments of the P. sativum phytoene desaturase (PDS), LEAFY (LFY) and KORRIGAN1 (KOR1) homologues were inserted into the PEBV RNA2 vector, replacing the genes required for nematode transmission. Pisum sativum inoculated with PEBV carrying a fragment of PsPDS developed characteristic photo‐bleached leaves and this phenotype was associated with a significant reduction in PsPDS mRNA. The P. sativum homologue of LFY is known as UNIFOLIATA (UNI). Plants inoculated with PEBV carrying a fragment of UNI developed distorted flowers and leaves with modified architecture, which are also observed in UNI‐mutants. In Arabidopsis thaliana, the KOR1‐mutant is characterized by an extreme dwarf phenotype. Pisum sativum plants inoculated with PEBV carrying a fragment of PsKOR1 displayed a significant reduction in height and inhibition of root growth. The PEBV VIGS vector did not affect the ability of P. sativum to flower, set seeds, and form nodules characteristic of symbiosis with rhizobium. These results suggest that the PEBV vector can be applied to functional genomics in a legume species to study genes involved in a wide range of biological processes.
The mitochondrion – an organelle commonly involved in programmed cell death in Arabidopsis thaliana
Authors: Yao Nan , Eisfelder Bartholomew J. , Marvin James , Greenberg Jean T.
Published: December 2004

Plant cells undergoing programmed cell death (PCD) at late stages typically show chromatin condensation and endonucleolytic cleavage prior to obvious membrane or organelle ultrastructural changes. To investigate possible early PCD‐associated events, we used microscopic observations and flow cytometry to quantitate mitochondrial membrane potential (ΔΨm) changes during PCD at the single cell and population levels using Arabidopsis protoplasts. A ΔΨm loss was commonly induced early during plant PCD and was important for PCD execution, as evidenced by the concomitant reduction of the change in ΔΨm and PCD by cyclosporin A, which inhibits mitochondrial permeability transition pores in animal cells. ΔΨm loss occurred prior to nuclear morphological changes and was only associated with mitochondrial cytochrome c release (an apoptotic trigger in animals) in response to one of three PCD elicitors. Three different stimuli in wild type implicated ΔΨm changes in PCD: ceramide, protoporphyrin IX, and the hypersensitive response elicitor AvrRpt2. Additionally, the behavior of the conditional ectopic cell death mutant accelerated cell death2 and ACD2‐overproducing plants also implicated ΔΨm alteration as key for PCD execution. Because ACD2 is largely a chloroplast component in mature plants, the observation that the cell death in acd2 mutants requires changes in mitochondrial functions implicates communication between chloroplasts and mitochondria in mediating PCD activation. We suggest that ΔΨm loss is a common early marker in plant PCD, similar to what has been documented in animals. However, unlike in animal cells, in plant cells, mitochondrial cytochrome c release is not an obligatory step in PCD control.
Drug resistance in Plasmodium falciparum from the Chittagong Hill Tracts, Bangladesh
Authors: Van Den Broek Ingrid V. F. , Van Der Wardt Sophia , Talukder Lenin , Chakma Swarupa , Brockman Alan , Nair Shalini , Anderson Tim C.
Published: December 2004

Objective  To assess the efficacy of antimalarial treatment and molecular markers of Plasmodium falciparum resistance in the Chittagong Hill Tracts of Bangladesh.

Methods  A total of 203 patients infected with P. falciparum were treated with quinine 3 days plus sulphadoxine/pyrimethamine (SP) combination therapy, and followed up during a 4‐week period. Blood samples collected before treatment were genotyped for parasite mutations related to chloroquine (pfcrt and pfmdr1 genes) or SP resistance (dhfr and dhps).

Results  Of 186 patients who completed follow‐up, 32 patients (17.2%) failed to clear parasitaemia or became positive again within 28 days after treatment. Recurring parasitaemia was related to age (χ2 = 4.8, P < 0.05) and parasite rates on admission (t = 3.1, P < 0.01). PCR analysis showed that some of these cases were novel infections. The adjusted recrudescence rate was 12.9% (95% CI 8.1–17.7) overall, and 16.6% (95% CI 3.5–29.7), 15.5% (95% CI 8.3–22.7) and 6.9% (95% CI 0.4–13.4) in three age groups (<5 years, 5–14, ≥15). The majority of infections carried mutations associated with chloroquine resistance: 94% at pfcrt and 70% at pfmdr. Sp‐resistant genotypes were also frequent: 99% and 73% of parasites carried two or more mutations at dhfr and dhps, respectively. The frequency of alleles at dhfr, dhps and pfmdr was similar in cases that were successfully treated and those that recrudesced.

Conclusions  The clinical trial showed that quinine 3‐days combined to SP is still relatively effective in the Chittagong Hill Tracts. However, if this regimen is continued to be widely used, further development of SP resistance and reduced quinine sensitivity are to be expected. The genotyping results suggest that neither chloroquine nor SP can be considered a reliable treatment for P. falciparum malaria any longer in this area of Bangladesh.
Measuring the efficacy of insecticide treated bednets: the use of DNA fingerprinting to increase the accuracy of personal protection estimates in Tanzania
Authors: Soremekun Seyi , Maxwell Caroline , Zuwakuu Martin , Chen Cynthia , Michael Edwin , Curtis Christopher
Published: December 2004

Insecticide‐treated nets have proved successful in the prevention of malaria as a result of both the personal protection with which they provide the sleeper and also the ‘mass effect’ on the local mosquito population when they are used on a community‐wide basis. Personal protection estimates are normally based on comparisons of the numbers of bloodfed mosquitoes found in rooms with and without nets, however it seemed possible that a number of those mosquitoes may not have fed on the occupants of the rooms in which they were found but had entered after feeding elsewhere. To address this possible source of error, we used an 8‐locus microsatellite system to identify the source of bloodmeals of Anopheles gambiaes.l. and A. funestus mosquitoes collected in rooms and window traps in Tanzanian villages with and without nets treated with alphacypermethrin. DNA fingerprints were produced from blood samples taken from people who had slept in these rooms and were matched to fingerprints obtained from the mosquito bloodmeals. We were able to type successfully over 90% of the bloodmeals collected and found that proportions of bloodfed mosquitoes that had fed on occupants of the rooms in which they were found were high and only slightly greater in villages without treated nets than those with them (95% and 88%, respectively). When these percentages were used to adjust estimates of personal protection, it was found that the error due to mosquitoes not feeding in the rooms in which they were collected is negligible.
The prevalence, intensities and risk factors associated with geohelminth infection in tea‐growing communities of Assam, India
Authors: Traub Rebecca J. , Robertson Ian D. , Irwin Peter , Mencke Norbert , Andrew Thompson R. C.
Published: December 2004

Objective  To determine the prevalence, intensity and associated risk factors for infection with Ascaris, hookworms and Trichuris in three tea‐growing communities in Assam, India.

Methods  Single faecal samples were collected from 328 individuals and subjected to centrifugal floatation and the Kato Katz quantitation technique and prevalence and intensities of infection with each parasite calculated. Associations between parasite prevalence, intensity and host and environmental factors were then made using both univariate and multivariate analysis.

Results  The overall prevalence of Ascaris was 38% [95% confidence interval (CI): 33, 43], and the individual prevalence of hookworm and Trichuris was 43% (95% CI: 38, 49). The strongest predictors for the intensity of one or more geohelminths using multiple regression (P ≤ 0.10) were socioeconomic status, age, household crowding, level of education, religion, use of footwear when outdoors, defecation practices, pig ownership and water source.

Conclusion  A universal blanket treatment with broad‐spectrum anthelmintics together with promotion of scholastic and health education and improvements in sanitation is recommended for helminth control in the communities under study.
Asking semi‐literate adolescents about sexual behaviour: the validity of assisted self‐completion questionnaire (ASCQ) data in rural Tanzania
Authors: Plummer Mary L. , Wight Daniel , Ross David A. , Balira Rebecca , Anemona Alessandra , Todd Jim , Salamba Zachayo , Obasi Angela I. N. , Grosskurth Heiner , Changalunga John , Hayes Richard J.
Published: December 2004

Objectives  To develop and test a sexual behaviour survey method for semi‐literate populations, combining the privacy of a self‐completion questionnaire (SCQ) with the clarity of a face‐to‐face questionnaire (FFQ).

Methods  In 1998, 6079 Tanzanian primary school students (mean age 15.1 years) were surveyed using an innovative assisted self‐completion questionnaire (ASCQ). The format of the questionnaire was simple, all responses were closed, and conceptually complex questions such as those involving ranking or multiple answers were avoided. The ASCQ was administered to groups of 20 by a research assistant who read questions and answers aloud in two languages so pupils could tick or write responses independently. A total of 4958 of respondents from the 1998 ASCQ Cohort also participated in a 1998 FFQ interview and, in 2000, 4424 again completed an ASCQ.

Results  In the 1998 ASCQ survey, 55.0% of males and 21.1% of females reported they had had vaginal intercourse, of whom 71.5% and 66.0%, respectively reported their first sexual relationship lasted for a week or less, and 49.5% and 59.6%, respectively reported they had had sex in the last 4 weeks. After adjustment for age, reported sex was associated with alcohol use in both males (OR = 1.57) and females (OR = 1.69), earning money for males (OR = 1.32) and not living with a mother for females (OR = 0.77). The vast majority of respondents did not appear to have difficulty completing the ASCQ, but 7.4% of 1998 respondents and 2.9% of 2000 respondents selected all first or all last answers in a section for which this was inconsistent. This bias was associated with female, less educated and more geographically remote respondents. Of those respondents who reported sex in the 1998 ASCQ survey, 32.1% reported fewer total partners in the 2000 ASCQ survey, 25.2% reported having had sex fewer times than originally reported, and 61.9% of those who reported having used a condom in 1998 reported never having used one in 2000. While the proportions reporting sex were very similar in the 1998 ASCQ and FFQ surveys, 37.9% of males and 59.2% of females reporting sex only did so on one of the two questionnaires. Higher proportions of respondents reported sensitive information in the ASCQ than the FFQ, although in some cases this may have related to answer order bias.

Conclusion  The results suggest that an ASCQ may be useful in assessing sexual behaviour in African adolescents, particularly for older, male and/or educated respondents. However, triangulation with data from other surveys raises questions about the validity of self‐reported sexual behaviour in general.
Efficacité de combinaisons thérapeutiques avec des dérivés de l'artémisinine dans le traitement de l'accès palustre non‐complique au Burundi
Authors: Ndayiragije Athanase , Niyungeko Déo , Karenzo Jeanne , Niyungeko Ernest , Barutwanayo Marianne , Ciza Alphonse , Bosman Andrea , Moyou‐Somo Roger , Nahimana Adélaïde , Nyarushatsi Jean Paul , Barihuta Tharcisse , Mizero Liévin , Ndaruhutse Jérome , Delacollette Charles , Ringwald Pascal , Kamana Jean
Published: December 2004

Confrontées au problème de la résistance à la chloroquine et à la sulfadoxine‐pyriméthamine, les autorités du Burundi ont choisi d’étudier l'efficacité de deux combinaisons à base de dérivés de l'artemisinine, d'une part l'association fixe artéméther‐luméfantrine et d'autre part l'association non‐fixe artésunate + amodiaquine. Les combinaisons médicamenteuses ont étéétudiées dans 2 sites représentatifs du pays à Kigobe, un site urbain situé dans la périphérie de Bujumbura et à Buhiga en milieu rural. L’étude a été menée selon le protocole de l'OMS modifié en 2001 d'octobre 2001 à novembre 2002. Au total 295 enfants ont été recrutés, 153 enfants ont été traités avec l'association artésunate + amodiaquine (77 à Buhiga et 76 à Kigobe) et 142 patients avec l'association artéméther‐luméfantrine (64 à Buhiga et 78 à Kigobe). Parmi les 295 enfants, 290 ont été suivis jusqu’à J14. Sur les 149 enfants qui ont reçu la combinaison artésunate + amodiaquine, 142 (95,3%, IC95%: 91,9–98,7%) ont présenté une réponse clinique et parasitologique adéquate, 5 (3,3%) un échec parasitologique tardif, 1 (0,7%) un échec clinique tardif et 1 (0,7%) un échec thérapeutique précoce. Sur les 141 enfants ayant reçu la combinaison artéméther‐luméfantrine, 140 (99,3%, CI95%: 97,9–100%) ont présenté une réponse clinique et parasitologique adéquate et 1 (0,7%) un échec parasitologique tardif à Buhiga. Les effets secondaires étaient similaires dans les deux groupes de traitement, excepté les vomissements qui étaient significativement plus fréquents à J1 et J2 dans le groupe artésunate + amodiaquine. Les deux traitements ont eu une action sur le nombre de porteurs de gamétocytes sans obtenir une disparition totale chez tous les malades. Au cours d'une réunion de consensus, le Ministère de la Santé Publique du Burundi a choisi de sélectionner la combinaison thérapeutique artésunate + amodiaquine comme première ligne de traitement du paludisme non‐compliqué au Burundi y compris au cours des épidémies.
Bibliometric analysis of research published in Tropical Medicine and International Health 1996–2003
Authors: Glover Steven W. , Bowen Sarah L.
Published: December 2004

We examine the bibliometric profile of Tropical Medicine and International Health using the subjects of the articles published and the geographical distribution of the authors. The most common subject areas of papers published during 1996–2003 are highlighted, and the most cited papers indicated.
Predominance of lineage I among Trypanosoma cruzi isolates from Venezuelan patients with different clinical profiles of acute Chagas’ disease
Authors: Añez Nestor , Crisante Gladys , Silva Flavia Maia da , Rojas Agustina , Carrasco Hugo , Umezawa Eufrosina S. , Stolf Anna Maria S. , Ramírez Jose Luis , Teixeira Marta M. G.
Published: December 2004

Trypanosoma cruzi isolates from 23 acute chagasic patients from localities of Western Venezuela (state of Barinas) where Chagas’ disease is endemic were typed using ribosomal and mini‐exon gene markers. Results showed that isolates of the two major phylogenetic lineages, T. cruzi I and T. cruzi II, were isolated from these patients. Six isolates (26%) were typed as T. cruzi II and 17 (74%) as belonging to T. cruzi lineage I. Analysis of random amplified polymorphic DNA (RAPD) patterns confirmed these two groups of isolates, but did not disclose significant genetic intra‐lineage polymorphism. Patients infected by both T. cruzi I or T. cruzi II showed different clinical profiles presenting highly variable signs and symptoms of acute phase of Chagas’ disease ranging from totally asymptomatic to severe heart failure. The predominance of T. cruzi I human isolates in Venezuela allied to the higher prevalence of severe symptoms of Chagas’ disease (heart failure) in patients infected by this lineage do not corroborate an innocuousness of T. cruzi I infection to humans. To our knowledge, this is the first study describing predominance of T. cruzi lineage I in a large number of acute chagasic patients with distinct and well‐characterized clinical profiles.
Exploring 30 years of malaria case data in KwaZulu‐Natal, South Africa: Part II. The impact of non‐climatic factors
Authors: Craig M. H. , Kleinschmidt I. , Le Sueur D. , Sharp B. L.
Published: December 2004

Malaria transmission is a multifactorial phenomenon. Climate is a major limiting factor in the spatial and temporal distribution of malaria, but many non‐climatic factors may alter or override the effect of climate. Thirty years of monthly malaria incidence data from KwaZulu‐Natal province, South Africa, reveal strong medium and long‐term trends, which were not present in the climate data. This paper explores various non‐climatic factors that may have contributed towards the observed trends. The development of antimalarial drug resistance, available information on human immunodeficiency virus (HIV) prevalence, cross‐border people movements, agricultural activities, emergence of insecticide resistance and the case reporting system are reviewed and their potential effect on malaria transmission examined. Single‐variable linear regression analysis showed significant association between seasonal case totals (log‐transformed) and the measured level of drug resistance (log‐transformed) (r2 = 0.558, n = 10, P = 0.013) as well as relative measures of HIV infection since 1990 (r2 = 0.846, n = 11, P = 0.001). The other factors appear to have affected the level of malaria transmission at certain periods and to some degree. The importance of surveillance and inclusion of non‐climatic variables in analysis of malaria data is demonstrated.
High‐throughput genotyping of human platelet antigens using the 5′‐nuclease assay and minor groove binder probe technology
Authors: Higgins M. , Hughes A. , Buzzacott N. , Lown J.
Published: December 2004

Background and Objectives  Human platelet antigen (HPA) genotyping is a valuable tool for typing platelets to assist in the management of alloimmunized thrombocytopenic patients. We describe, for the first time, 5′ nuclease assays (NA) to genotype HPA‐5 and ‐15, and improved 5′‐NA to genotype HPA‐1, ‐2 and ‐3, by utilizing minor groove binder (MGB) and non‐fluorescent quencher (NFQ) technology. Superior probe specificity and fluorescent performance is attained through MGB‐NFQ probe modifications compared with previous 5′‐NA designs.

Materials and Methods  Primers and dye‐labelled MGB‐NFQ probes were designed and synthesized to detect the single nucleotide polymorphism responsible for each HPA‐1, ‐2, ‐3, ‐5 and ‐15. One‐hundred blood samples were tested for the combinations of HPA genotypes 1, 2, 3 and 5 by our traditional sequence‐specific primer–polymerase chain reaction (SSP–PCR) method, and 41 blood samples were tested for HPA‐15 by SSP–PCR at an external laboratory. These results were then compared with those obtained by using the new 5′‐NA.

Results  There was complete concordance of results for all samples tested by SSP–PCR and 5′‐NA. The 5′‐NA offers distinct advantages over non‐fluorescent genotyping methods. DNA amplification and allele discrimination occurs in a single closed tube for each antigen with no post‐PCR manipulation required. This minimizes the risk of cross‐contamination and mislabelling of samples, as well as making the assay less time‐consuming to perform. In comparison with other fluorescent assays, the 5′‐NA has the highest sample throughput, resulting from the use of a 96‐well platform, identical cycling conditions for all assays and the potential for automation.

Conclusions  Genotyping for HPA‐1, ‐2, ‐3, ‐5, and ‐15 by the 5′‐NA is suitable for routine analysis. The latest 5′‐NA design, using MGB probe technology, ensures superior detection of all alleles and is the most versatile fluorescent assay, ideal for both urgent clinical samples and large‐scale screening programs.
The Relations among Observational, Physiological, and Self‐Report Measures of Children's Anger
Authors: Hubbard Julie A. , Parker Elizabeth H. , Ramsden Sally R. , Flanagan Kelly D. , Relyea Nicole , Dearing Karen F. , Smithmyer Catherine M. , Simons Robert F. , Hyde Christopher T.
Published: December 2004

Our first goal was to examine the relations among observational, physiological, and self‐report measures of children's anger. Our second goal was to investigate whether these relations varied by reactive or proactive aggression. Children (272 second‐grade boys and girls) participated in a procedure in which they lost a game and prize to a confederate who cheated. Skin conductance reactivity, heart rate reactivity, self‐reports of anger, angry facial expressions, and angry nonverbal behaviors were measured for each turn of the game. We used multi‐level regressions to calculate the relations among the 10 pairs of the five anger variables over the course of the game. Six of the 10 pairs of anger variables were positively related. These findings suggest that measuring children's anger using any one approach may not capture the full complexity of children's overall experience and expression of anger. Furthermore, three of the 10 relations were stronger at higher levels of reactive aggression, although none varied by proactive aggression. These findings suggest that reactive aggression is related to greater cohesiveness in the experience and expression of anger than is proactive aggression.
Mothers’ Regulation Strategies in Response to Toddlers’ Affect: Links to Later Emotion Self‐Regulation
Authors: Spinrad Tracy L. , Stifter Cynthia A. , Donelan‐McCall Nancy , Turner Laura
Published: December 2004

Recently, there has been a great deal of research on the socialization of children's emotions and self‐regulation. In the present study, the specific strategies that mothers use to help their young children regulate their emotional responses were examined using a longitudinal design. Forty‐three mother–toddler pairs were observed when toddlers were both 18 and 30 months of age, and mothers’ attempts to regulate their toddlers’ emotions during several emotion‐eliciting tasks were transcribed from videotape. When the children were 5 years old, their responses to a disappointment task were observed. Results indicated a relation between mothers’ regulation strategies in toddlerhood and children's facial and behavioral responses to the disappointment task measured at 5 years of age. Specifically, mothers’ use of regulation strategies at 30 months, but not at 18 months, was positively related to children's appropriate emotional displays in response to disappointment. Moreover, the specific types of strategies that mothers used had differential associations to children's responses to disappointment. Findings are discussed in terms of the potentially important role of mothers’ behaviors in the development of children's emotion self‐regulation.
A Short‐term Longitudinal Study of Children's Playground Games in Primary School: Implications for Adjustment to School and Social Adjustment in the USA and the UK
Authors: Pellegrini Anthony D. , Blatchford Peter , Kato Kentaro , Baines Ed
Published: December 2004

In this study we described development of boys’ and girls’ games on the school playground at recess as they progressed across the first year of primary school in London (UK) and Minneapolis (USA) in order to document age‐related trends in games during this period and to examine the predictive role of games in children's adjustment to school and more general social adjustment. Consistent with expectations, games accounted for a significant portion of children’s, but especially boys’, behavior during recess. American children played games more frequently, especially simple games, relative to English children. Consistent with expectation, the complexity of games increased, especially for boys, across the school year for children in both countries. We also examined the extent to which facility with games forecasted adjustment to school and social adjustment. Games predicted both, but only in the USA sample. Results are discussed in terms of games being an important developmental task for children of this age. Implications for future research and educational policy are discussed.
Repeated Screening with Inspection Error and No False Positive Results with Application to Pharmaceutical Pill Production
Authors: Gasparini Mauro , Nusser Harald , Eisele Jeffrey
Published: December 2004

Repeated screening is a 100% sampling inspection of a batch of items followed by removal of the defective items and further iterations of inspection and removal. The reason for repeating the inspection is that the detection of a defective item happens with probability p<1. A missed defective item is a false negative result. The no false positive result is contemplated in this paper, which is motivated by a problem coming from the production of pharmaceutical pills. Bayesian posterior distributions for the quality of the lot are obtained for the case of both p known and p unknown. Batch rejection and batch acceptance control limits for the number of defective items at subsequent iterations can then be calculated. Theoretical connections to the problem of estimating the number-of-trials parameter of a binomial distribution are drawn.
Signal transduction by G‐proteins, Rho‐kinase and protein phosphatase to smooth muscle and non‐muscle myosin II
Authors: Somlyo Andrew P. , Somlyo Avril V.
Published: December 2004

We here review mechanisms that can regulate the activity of myosin II, in smooth muscle and non‐muscle cells, by modulating the Ca2+ sensitivity of myosin regulatory light chain (RLC) phosphorylation. The major mechanism of Ca2+ sensitization of smooth muscle contraction and non‐muscle cell motility is through inhibition of the smooth muscle myosin phosphatase (MLCP) that dephosphorylates the RLC in smooth muscle and non‐muscle. The active, GTP‐bound form of the small GTPase RhoA activates a serine/threonine kinase, Rho‐kinase, that phosphorylates the regulatory subunit of MLCP and inhibits phosphatase activity. G‐protein‐coupled release of arachidonic acid may also contribute to inhibition of MLCP acting, at least in part, through the Rho/Rho‐kinase pathway. Protein kinase C(s) activated by phorbol esters and diacylglycerol can also inhibit MLCP by phosphorylating and thereby activating CPI‐17, an inhibitor of its catalytic subunit; this mechanism is independent of the Rho/Rho‐kinase pathway and plays only a minor, transient role in the G‐protein‐coupled mechanism of Ca2+ sensitization. Ca2+ sensitization by the Rho/Rho‐kinase pathway contributes to the tonic phase of agonist‐induced contraction in smooth muscle, and abnormally increased activation of myosin II by this mechanism is thought to play a role in diseases such as high blood pressure and cancer cell metastasis.
Differentiated practice, patient‐oriented care and quality of work in a hospital in the Netherlands
Authors: Boumans Nicolle P. G. , Landeweerd Jan A. , Visser Mildred
Published: December 2004

This article describes a quantitative study of the relationship between differentiated practice on the one side and patient‐oriented care and quality of work on the other. Nursing wards where differentiated practice has been implemented (intervention group) have been compared with wards where differentiated practice has not been implemented (reference group). The research variables with regard to differentiated practice, patient‐oriented care and quality of work have been measured by questionnaires. Subjects were 68 nurses and six supervisors from six nursing wards from one hospital. The results show that the extent to which differentiated practice had been implemented varied between the wards. With regard to patient‐oriented care differences have been found between the intervention and reference group on the variables patient assignment and use of the nursing process, but not on the variables of tasks and communication. Concerning quality of work, differences have been found on: social support from the supervisor, social–emotional leadership and health complaints. Rank order correlations between differentiated practice and patient‐oriented care and between differentiated practice and quality of work were not significant. Practical implications with regard to the use of differentiated practice and implications for further research are discussed.