On species delimitation, hybridization and population structure of cassava whitefly in Africa.

The Bemisia cassava whitefly complex includes species that cause severe crop damage through vectoring cassava viruses in eastern Africa. Currently, this whitefly complex is divided into species and subgroups (SG) based on very limited molecular markers that do not allow clear definition of species and population structure. Based on 14,358 genome-wide SNPs from 62 Bemisia cassava whitefly individuals belonging to sub-Saharan African species (SSA1, SSA2 and SSA4), and using a well-curated mtCOI gene database, we show clear incongruities in previous taxonomic approaches underpinned by effects from pseudogenes. We show that the SSA4 species is nested within SSA2, and that populations of the SSA1 species comprise well-defined south-eastern (Madagascar, Tanzania) and north-western (Nigeria, Democratic Republic of Congo, Burundi) putative sub-species. Signatures of allopatric incipient speciation, and the presence of a 'hybrid zone' separating the two putative sub-species were also detected. These findings provide insights into the evolution and molecular ecology of a highly cryptic hemipteran insect complex in African, and allow the systematic use of genomic data to be incorporated in the development of management strategies for this cassava pest.

Cassava whitefly, Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) in East African farming landscapes: a review of the factors determining abundance.

Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) is a pest species complex that causes widespread damage to cassava, a staple food crop for millions of households in East Africa. Species in the complex cause direct feeding damage to cassava and are the vectors of multiple plant viruses. Whilst significant work has gone into developing virus-resistant cassava cultivars, there has been little research effort aimed at understanding the ecology of these insect vectors. Here we assess critically the knowledge base relating to factors that may lead to high population densities of sub-Saharan African (SSA) B. tabaci species in cassava production landscapes of East Africa. We focus first on empirical studies that have examined biotic or abiotic factors that may lead to high populations. We then identify knowledge gaps that need to be filled to deliver sustainable management solutions. We found that whilst many hypotheses have been put forward to explain the increases in abundance witnessed since the early 1990s, there are little published data and these tend to have been collected in a piecemeal manner. The most critical knowledge gaps identified were: (i) understanding how cassava cultivars and alternative host plants impact population dynamics and natural enemies; (ii) the impact of natural enemies in terms of reducing the frequency of outbreaks and (iii) the use and management of insecticides to delay the development of resistance. In addition, there are several fundamental methodologies that need to be developed and deployed in East Africa to address some of the more challenging knowledge gaps.

Understanding reconstructed Dante spectra using high resolution spectroscopy.

The Dante is an 18 channel filtered diode array used at the National Ignition Facility (NIF) to measure the spectrally and temporally resolved radiation flux between 50 eV and 20 keV from various targets. The absolute flux is determined from the radiometric calibration of the x-ray diodes, filters, and mirrors and a reconstruction algorithm applied to the recorded voltages from each channel. The reconstructed spectra are very low resolution with features consistent with the instrument response and are not necessarily consistent with the spectral emission features from the plasma. Errors may exist between the reconstructed spectra and the actual emission features due to assumptions in the algorithm. Recently, a high resolution convex crystal spectrometer, VIRGIL, has been installed at NIF with the same line of sight as the Dante. Spectra from L-shell Ag and Xe have been recorded by both VIRGIL and Dante. Comparisons of these two spectroscopic measurements yield insights into the accuracy of the Dante reconstructions.

The NIF x-ray spectrometer calibration campaign at Omega.

The calibration campaign of the National Ignition Facility X-ray Spectrometer (NXS) was carried out at the Omega laser facility. Spherically symmetric, laser-driven, millimeter-scale x-ray sources of K-shell and L-shell emission from various mid-Z elements were designed for the 2-18 keV energy range of the NXS. The absolute spectral brightness was measured by two calibrated spectrometers. We compare the measured performance of the target design to radiation hydrodynamics simulations.

Demonstration of a 13-keV Kr K-shell x-ray source at the National Ignition Facility.

We report 3% conversion efficiency of laser energy into Kr K-shell (≈13 keV) radiation, consistent with theoretical predictions. This is ≈10× greater than previous work. The emission was produced from a 4.1-mm-diameter, 4-mm-tall gas pipe target filled with 1.2 or 1.5 atm of Kr gas. 160 of the National Ignition Facility laser beams deposited ≈700 kJ of 3ω light into the target in an ≈140 TW, 5.0-ns-duration square pulse. The Dante diagnostics measured ≈5 TW into 4π solid angle of ≥12 keV x rays for ≈4 ns, which includes both continuum emission and flux in the Kr He_{α} line at 13 keV.

Quantitative trait loci linked to thalamus and cortex gray matter volumes in BXD recombinant inbred mice.

To investigate whether there are separate or shared genetic influences on the development of the thalamus and cerebral cortex, we identified quantitative trait loci (QTLs) for relevant structural volumes in BXD recombinant inbred (RI) strains of mice. In 34 BXD RI strains and two parental strains (C57BL/6J and DBA/2J), we measured the volumes of the entire thalamus and cortex gray matter using point counting and Cavalieri's rule. Heritability was calculated using analysis of variance (ANOVA), and QTL analysis was carried out using WebQTL (http://www.genenetwork.org). The heritability of thalamus volume was 36%, and three suggestive QTLs for thalamus volume were identified on chromosomes 10, 11 and 16. The heritability of cortical gray matter was 43%, and four suggestive QTLs for cortex gray matter volume were identified on chromosomes 2, 8, 16 and 19. The genetic correlation between thalamus and cortex gray matter volumes was 0.64. Also, a single QTL on chromosome 16 (D16Mit100) was identified for thalamus volume, cortex gray matter volume and Morris water maze search-time preference (r=0.71). These results suggest that there are separate and shared genetic influences on the development of the thalamus and cerebral cortex.

PCR-based detection and partial genome sequencing indicate high genetic diversity in Bangladeshi begomoviruses and their whitefly vector, Bemisia tabaci.

The population diversity of Bangladeshi begomoviruses and their vector, Bemisia tabaci was analysed by PCR-based detection and partial genome sequencing. B. tabaci adults and plants expressing symptoms of virus infection were collected from locations representing diverse agro-ecological regions of the country. Universal and species-specific primers were used to detect begomoviruses in seven crops (chilli, okra, papaya, pumpkin, sponge gourd, tomato and yardlong bean) and two common weeds (Ageratum conyzoides and Croton bonplandianum). At least five distinct species of tomato leaf curl viruses infected tomato and other host-plants. Phylogenetic analyses of their nucleotide sequences ( approximately 530 bases) from the intergenic region and capsid protein of DNA-A indicated the existence of five distinct clusters of begomoviruses. Begomoviruses infecting tomato, chilli and dolichos have been reported previously, and those infecting Ageratum, Croton, okra, papaya, pumpkin and yardlong bean are described for the first time. Phylogenetic analyses based on mitochondrial cytochrome oxidase I gene sequences of 21 B. tabaci from Bangladesh and other reference sequences grouped them into at least two independent clusters. Some sequences from different countries, e.g., Bangladesh, China, India, Nepal, Pakistan and Thailand were almost identical while others collected from plants within the same field diverged by as much as 15%, indicating high diversity even at the local level. None of the B. tabaci from Bangladesh grouped with the reference B- and Q-biotype sequences, thus these two aggressive biotypes were apparently absent from Bangladesh.

Re-emergence of Cassava Brown Streak Disease in Uganda.

During November 2004, veinal chlorosis on mature cassava leaves, typical of cassava brown streak disease (CBSD), was observed at Mukono in central Uganda. Five out of 11 cultivars at the site showed CBSD symptoms (incidence range 4 to 64%). In a survey of farmers' fields, CBSD was observed in Wakiso and Mukono districts. Incidence of cassava mosaic disease was also recorded and averaged 60% for landraces (range 16.7 to 100%) and 20% for resistant varieties (range 0 to 65%). Leaf samples of plants with CBSD symptoms produced an amplicon of 222 bp using reverse transcription-polymerase chain reaction with primers that amplify a fragment of the coat protein (CP) gene of Cassava brown streak virus. Sequence comparisons based on the amplified CP gene fragment indicated that the isolates have 77 to 82.9% nucleotide and 43.9 to 56.8% amino acid identity with those from Mozambique and Tanzania. There was 95.9 to 99.5% nucleotide and 85.1 to 90.5% amino acid identity among the Ugandan isolates. These results confirm the re-emergence of CBSD in Uganda after it was first observed in the 1930s in cassava introduced from Tanzania and controlled by eradication. Prior to this report, CBSD was known to be restricted to the coastal lowlands of East Africa.

Predicting the thermal inactivation of bacteria in a solid matrix: simulation studies on the relative effects of microbial thermal resistance parameters and process conditions.

A combined mathematical model for predicting heat penetration and microbial inactivation in a solid body heated by conduction was tested experimentally by inoculating agar cylinders with Salmonella typhimurium or Enterococcus faecium and heating in a water bath. Regions of growth where bacteria had survived after heating were measured by image analysis and compared with model predictions. Visualisation of the regions of growth was improved by incorporating chromogenic metabolic indicators into the agar. Preliminary tests established that the model performed satisfactorily with both test organisms and with cylinders of different diameter. The model was then used in simulation studies in which the parameters D, z, inoculum size, cylinder diameter and heating temperature were systematically varied. These simulations showed that the biological variables D, z and inoculum size had a relatively small effect on the time needed to eliminate bacteria at the cylinder axis in comparison with the physical variables heating temperature and cylinder diameter, which had a much greater relative effect.

Direct observation of the alpha-epsilon transition in shock-compressed iron via nanosecond x-ray diffraction.

In situ x-ray diffraction studies of iron under shock conditions confirm unambiguously a phase change from the bcc (alpha) to hcp (epsilon) structure. Previous identification of this transition in shock-loaded iron has been inferred from the correlation between shock-wave-profile analyses and static high-pressure x-ray measurements. This correlation is intrinsically limited because dynamic loading can markedly affect the structural modifications of solids. The in situ measurements are consistent with a uniaxial collapse along the [001] direction and shuffling of alternate (110) planes of atoms, and are in good agreement with large-scale nonequilibrium molecular dynamics simulations.

Pall eBDS: an enhanced bacterial detection system for screening platelet concentrates.

Bacterial contamination of blood components remains a significant problem in transfusion medicine. The Pall enhanced bacterial detection system (Pall eBDS) detects the presence of bacteria in leucodepleted platelet concentrates by measuring the reduction of oxygen in the sample, due to aerobic bacterial growth. Pooled platelet concentrates were spiked at 10 cfu mL(-1) with 10 organisms (one species per bag). Pall eBDS pouches were inoculated with the spiked platelet concentrates. After 24 and 30 h of incubation, the oxygen level was measured. A further set of pouches were taken from the inoculated platelet concentrates at 24 h. Incubation and reading intervals were as for the initial set of pouches. A sensitivity study was also performed comparing the Pall eBDS with the BacT/ALERT system. Spiking at 10 cfu mL(-1) and immediately sampling into Pall eBDS pouches resulted in 97.6 and 100% detection after an incubation period of 24 and 30 h, respectively. After 24 h of incubation of the spiked platelet concentrates and then sampling into Pall eBDS pouches, 99.1% detection was obtained after incubation for both 24 and 30 h. The sensitivity of the Pall eBDS and BacT/ALERT is similar and in the order of 1 cfu mL(-1). Implementation of either BacT/ALERT or Pall eBDS for routine screening of platelet concentrates has the potential to further increase the safety of the blood supply.

Use of a solid-phase fluorescent cytometric technique for the detection of bacteria in platelet concentrates.

Blood services worldwide are now striving to reduce the risk of transmission of bacteria by transfusion. The BacT/ALERT microbial detection system (bioMerieux, Basingstoke, Hants, UK) is currently regarded as the 'gold standard' for bacterial screening of platelet concentrates. The BacT/ALERT is a culture system and will not generate an 'instant' (within 2 h) determination. We report on the Scansystem (Hemosystem, Marseille, France), a solid-phase fluorescent cytometric technique, which enables the rapid detection of bacteria (within 90 min) in platelet concentrates. The study was performed in two parts - one involving the routine screening of platelet concentrates and the other determining the sensitivity of the system. In both arms of the study, the BacT/ALERT was used for comparative purposes. In total, 900 platelet concentrates were screened (63 apheresis and 837 buffy coat pooled). No bacteria were detected in any of the platelet concentrates tested by means of either the Scansystem or the BacT/ALERT. The sensitivity of the Scansystem was in the order of 10(3) cfu mL(-1). Escherichia coli and Staphylococcus aureus were detected by using the Scansystem at 1 cfu mL(-1). The BacT/ALERT detected all organisms tested (n = 6) at 1 cfu mL(-1). The Scansystem offers a sensitive alternative technology to bacterial culture, with the benefit of a rapid test time.

Incidence of lower thoracic ligamentum flavum midline gaps.

BACKGROUND: Lower thoracic epidural anaesthesia and analgesia (EDA) has gained increasing importance in perioperative pain therapy. The loss-of-resistance technique used to identify the epidural space is thought to rely on the penetration of the ligamentum flavum. Investigations at the cervical and lumbar regions have demonstrated that the ligamentum flavum frequently exhibits incomplete fusion at different vertebral levels. Therefore, the aim of this study was to directly investigate the incidence of lower thoracic ligamentum flavum midline gaps in embalmed cadavers. METHODS: Vertebral column specimens were obtained from 47 human cadavers. Ligamentum flavum midline gaps were recorded between the vertebral levels T6 and L1. RESULTS: The incidence of midline gaps/number of viable specimens at the following levels was: T6-7: 2/45 (4.4%), T7-8: 1/47 (2.1%), T8-9: 2/45 (4.4%), T9-10: 7/39 (17.9%), T10-11: 12/34 (35.2%), T11-12: 10/35 (28.5%), T12/L1: 6/38 (15.8%). CONCLUSIONS: In the present study we have determined the frequency of lower thoracic ligamentum flavum midline gaps. Gaps are less frequent than at cervical levels, but more frequent than at lumbar levels. Peak incidence was found in the region between T10 and T12. Using a strict midline approach, one cannot therefore rely on the ligamentum flavum to impede entering the epidural space in all patients.

[Evaluation of a cadaver workshop for education in regional anesthesia]. / Bewertung eines Anatomieworkshops für die Regionalanästhesieausbildung.

BACKGROUND: The aim of the present investigation was to survey former participants (n=869) of a cadaver workshop using a mail questionnaire to assess the demographic data and the impact of these courses on daily practice. METHODS: The deadline for acceptance of return mail was 60 days. Descriptive statistics were employed for analysis of results. RESULTS: The response rate was 36.7% and the course was judged to be recommendable by 98.2%. The average course attendant was board certified and had spent a mean time of 9+/-6 years in anesthesiology. The highest quality and degree of subsequent practicability in daily routine was attributed to peripheral nerve block training on cadavers. Of the course participants two-thirds performed regional anesthesia procedures more often following attendance. The majority of course attendants had to defray at least a part of the course fee themselves, and one-third was required to invest leisure time to attend. CONCLUSION: Attendance of a cadaver workshop increased knowledge of clinical anatomy and enhanced performance of regional anesthesia procedures. Courses of this format constitute a currently underestimated adjunct to contemporary regional anesthesia education.

Diversity and distribution of begomoviruses infecting tomato in India.

Leaf curl begomoviruses cause serious yield losses to Indian tomato crops. Total DNAs were extracted from leaves of 69 tomato plants and 34 weeds or neighbouring crops collected from all the major tomato producing areas of India. Eighty-one of the 103 samples were positive by PCRs using begomovirus genus-specific primers. Coat protein (CP) genes from 29 samples were PCR amplified, cloned and sequenced. Phylogenetic analyses of the CP sequences revealed five different tomato leaf curl begomovirus (TLCB) clusters each <88% identity to the others. Four clusters represented known Indian TLCBs, whereas one cluster contained sequences originating from Haryana State with most identity (89%) to the provisional Begomovirus species Croton yellow vein mosaic virus.Sixty-five begomovirus positive samples were characterised further by PCR with DNA-beta, DNA-B, four Indian TLCB species, PALIc1960/PARIv722 (universal begomovirus primers), and by sequencing. The majority of samples represented monopartite TLCBs associated with DNA-beta components. All four known TLCBs appeared to be present throughout India. TLCBs were also present in chilli, cowpea, okra and tobacco crops, as well as in some common weeds. Papaya leaf curl virus and Pepper leaf curl Bangladesh virus sequences were detected in tomato. Mixed begomovirus infections, a prerequisite for recombination, were evident in 13 samples.

Nucleotide sequencing, whitefly transmission, and screening tomato for resistance against two newly described begomoviruses in bangladesh.

ABSTRACT The molecular diversity of Tomato leaf curl viruses (ToLCVs), from the two main tomato growing areas of Jessore and Joydebpur, Bangladesh, was investigated. The viral DNA was amplified from tomato plants exhibiting mild and severe symptoms by polymerase chain reaction, and the complete genomes of the ToLCVs were sequenced. An isolate of the bipartite Tomato leaf curl New Delhi virus-Severe (ToLCNDV-Svr) was associated with the severe symptom phenotype from Jessore (ToLCNDV-Svr[Jes]). A previously undescribed monopartite virus, designated Tomato leaf curl Joydebpur virus-Mild (ToLCJV-Mld), was sequenced from plants showing mild symptoms. ToLCNDV-Svr[Jes] was most closely related to ToLCNDV-[Lucknow] at 95.7% nucleotide (nt) identity and Tomato leaf curl Gujarat virus-[Varanasi] at 90.6% nt identity, based on DNA-A and -B component sequences. ToLCJV-Mld was similar to Pepper leaf curl Bangladesh virus at 87.1% DNA-A nt identity. Identification of ToLCNDV-Svr[Jes] and ToLCJV-Mld was in addition to the previously described Tomato leaf curl Bangladesh virus, with which they shared 73.2 and 86.0% DNA-A nt identities, thus demonstrating the existence of at least three distinct viruses infecting tomato in Bangladesh. Nucleotide identities and placement in phylogenetic trees suggested that the three ToLCVs may have had different evolutionary pathways. The whitefly, Bemisia tabaci, transmitted the viruses of this study equally efficiently. Four tomato cultivars (TLB111, TLB130, TLB133, and TLB182) resistant/ tolerant to South Indian ToLCV were screened against the Bangladesh ToLCVs in 2003-04. Although challenged by diverse viruses and potentially mixed infections, disease incidence remained low (6 to 45%) in the resistant cultivars compared with local cultivars (68 to 100%).

First Report of Tomato leaf curl New Delhi virus Infecting Tomato in Bangladesh.

Tomato is an important cash crop for resource-poor farmers and accounts for 20% of the 2 million t of vegetables grown annually in Bangladesh. Tomato cultivation is affected by Tomato leaf curl virus (ToLCV), which can cause as much as 100% yield loss. Plants exhibiting typical ToLCV disease symptoms of yellowing, severe leaf curling, and stunting were collected at Jessore, Bangladesh during September 2003. The putative virus was transmitted from tomato to tomato by the whitefly Bemisia tabaci. In two separate experiments, 100% transmission was achieved by using 10 viruliferous B. tabaci adults for each of the 20 test plants that was confirmed by comparing the symptoms on test and virus source plants. Total DNAs were extracted from the symptomatic leaves, and the putative viral genomes were amplified by polymerase chain reaction by using the Deng A and B primers (1). Sequences generated from these primers were used to design virus-specific primers that were used to obtain complete viral sequences. Full-length DNA-A (2,740 nt; GenBank Accession No. AJ875157) and DNA-B (2,688 nt; GenBank Accession No. AJ875158) sequences of a bipartite Tomato leaf curl New Delhi virus from Jessore (ToLCNDV-[Jes]) were obtained, which were most similar to the corresponding sequences of ToLCNDV-(Lucknow) (GenBank Accession No. Y16421) at 95.7% and Tomato leaf curl Gujarat virus-(Varanasi) (Gen-Bank Accession No. AY190291) at 90.6% nt identities, respectively. DNA-A sequences had only 73.2% nt identity with the previously reported monopartite Tomato leaf curl Bangladesh virus (GenBank Accession No. AF188481) (2), confirming the occurrence of mono- and bipartite bego-moviruses in Bangladesh. The virus diversity poses a challenge for ToLCVD management in Bangladesh. References: (1) D. Deng et al. Ann. Appl. Biol. 125:327, 1994. (2) S. K. Green et al. Plant Dis. 85:1286, 2001.

A novel method for the detection of bacteria in platelet concentrates utilizing oxygen consumption as a marker for bacterial growth.

Bacterial transfusion-transmission remains a significant problem in transfusion medicine. Diversion and improved donor arm disinfection has been introduced by blood services to reduce bacterial transmissions. These interventions are not 100% effective and, therefore, there is still a requirement to screen blood donations, particularly platelet concentrates which are responsible for the majority of transmissions. Pall BDS, a novel bacterial testing system, detects the presence of bacteria in platelet concentrates by measuring the reduction in oxygen content associated with bacterial growth. Buffy coat-derived pooled platelet concentrates were spiked with 12 aerobic and two anaerobic organisms (one species per bag, n = 10) at 100-700 cfu mL(-1). Samples were taken into Pall BDS sample pouches and incubated for 0, 24, 30 and 48 h. An initial incubation was undertaken at 35 degrees C for 24 h and subsequent incubation was at 22 degrees C. At the end of the incubation period the oxygen content in the Pall BDS pouches was measured using a gas analyser. An oxygen content less than or equal to 19.5% was deemed to be positive. Pall BDS pouches tested positive in 80, 94 and 98% units spiked with aerobic bacteria at 24, 30 and 48 h, respectively. Anaerobic bacteria were not detected by the system. Positive BDS pouches contained 10(6) cfu mL(-1) or greater. The system was simple and easy to perform. Pall BDS has a closed sampling system which prevents exogenous contamination. This initial study indicates that the Pall BDS offers a practicable system for detecting bacteria present in leucodepleted platelet concentrates.

East African cassava mosaic Zanzibar virus - a recombinant begomovirus species with a mild phenotype.

Cassava plants exhibiting mild symptoms of cassava mosaic disease (CMD) were collected from Unguja island, Zanzibar. Cuttings grown from these plants in the glasshouse produced similar symptoms, which were milder than those caused by other known cassava mosaic geminiviruses (CMGs). The whitefly vector, Bemisia tabaci (Gennadius), transmitted the putative virus to 27.7% (n = 18) of target plants. Total DNA extracted from diseased leaves did not yield diagnostic PCR-bands using virus-specific primers to known CMGs. Degenerate primers, however, produced a diagnostic band indicating the presence of a begomovirus. Full-length DNA-A (2785 nucleotides) and DNA-B (2763 nucleotides) components were subsequently PCR-amplified, cloned and sequenced. Phylogenetic analyses of DNA-A and -B sequences showed that they were most similar to strains of East African cassava mosaic virus from Tanzania and Uganda at 83% and 86% nucleotide identities, respectively. The number and arrangement of open reading frames were similar to those of bipartite begomoviruses from the Old World. DNA-A was predicted to have recombined in the intergenic region (IR), AC1 and AC4 genes, and DNA-B in the IR. A maximum nucleotide identity of 83% in the DNA-A component with other sequenced begomoviruses, together with different biological properties allows this virus to be recognised as belonging to a new species named East African cassava mosaic Zanzibar virus (EACMZV).