bluuebunny/crossref_metadata_2025 huggingface dataset:
Garrod's fourth inborn error of metabolism solved by the identification of mutations causing pentosuria
Authors: Pierce Sarah B. , Spurrell Cailyn H. , Mandell Jessica B. , Lee Ming K. , Zeligson Sharon , Bereman Michael S. , Stray Sunday M. , Fokstuen Siv , MacCoss Michael J. , Levy-Lahad Ephrat , King Mary-Claire , Motulsky Arno G.
Published: November 2011

Pentosuria is one of four conditions hypothesized by Archibald Garrod in 1908 to be inborn errors of metabolism. Mutations responsible for the other three conditions (albinism, alkaptonuria, and cystinuria) have been identified, but the mutations responsible for pentosuria remained unknown. Pentosuria, which affects almost exclusively individuals of Ashkenazi Jewish ancestry, is characterized by high levels of the pentose sugar
l
-xylulose in blood and urine and deficiency of the enzyme
l
-xylulose reductase. The condition is autosomal-recessive and completely clinically benign, but in the early and mid-20th century attracted attention because it was often confused with diabetes mellitus and inappropriately treated with insulin. Persons with pentosuria were identified from records of Margaret Lasker, who studied the condition in the 1930s to 1960s. In the
DCXR
gene encoding
l
-xylulose reductase, we identified two mutations,
DCXR c.583ΔC
and
DCXR c.52(+1)G > A
, each predicted to lead to loss of enzyme activity. Of nine unrelated living pentosuric subjects, six were homozygous for
DCXR c.583ΔC
, one was homozygous for
DCXR c.52(+1)G > A
, and two were compound heterozygous for the two mutant alleles.
l
-Xylulose reductase was not detectable in protein lysates from subjects’ cells and high levels of xylulose were detected in their sera, confirming the relationship between the
DCXR
genotypes and the pentosuric phenotype. The combined frequency of the two mutant
DCXR
alleles in 1,067 Ashkenazi Jewish controls was 0.0173, suggesting a pentosuria frequency of approximately one in 3,300 in this population. Haplotype analysis indicated that the
DCXR c.52(+1)G > A
mutation arose more recently than the
DCXR c.583ΔC
mutation.
Characterization of dsRNA-induced pancreatitis model reveals the regulatory role of
IFN regulatory factor 2
(
Irf2
) in
trypsinogen5
gene transcription
Authors: Hayashi Hideki , Kohno Tomoko , Yasui Kiyoshi , Murota Hiroyuki , Kimura Tohru , Duncan Gordon S. , Nakashima Tomoki , Yamamoto Kazuo , Katayama Ichiro , Ma Yuhua , Chua Koon Jiew , Suematsu Takashi , Shimokawa Isao , Akira Shizuo , Kubo Yoshinao , Mak Tak Wah , Matsuyama Toshifumi
Published: November 2011

Mice deficient for
interferon regulatory factor
(
Irf
)
2
(
Irf2
−/−
mice) exhibit immunological abnormalities and cannot survive lymphocytic choriomeningitis virus infection. The pancreas of these animals is highly inflamed, a phenotype replicated by treatment with poly(I:C), a synthetic double-stranded RNA. Trypsinogen5 mRNA was constitutively up-regulated about 1,000-fold in
Irf2
−/−
mice compared with controls as assessed by quantitative RT-PCR. Further knockout of
IFNα/β receptor 1
(
Ifnar1
) abolished poly(I:C)-induced pancreatitis but had no effect on the constitutive up-regulation of
trypsinogen5
gene, indicating crucial type I IFN signaling to elicit the inflammation. Analysis of
Ifnar1
−/−
mice confirmed type I IFN-dependent transcriptional activation of dsRNA-sensing pattern recognition receptor genes
MDA5
,
RIG-I
, and
TLR3
, which induced poly(I:C)-dependent cell death in acinar cells in the absence of IRF2. We speculate that Trypsin5, the
trypsinogen5
gene product, leaking from dead acinar cells triggers a chain reaction leading to lethal pancreatitis in
Irf2
−/−
mice because it is resistant to a major endogenous trypsin inhibitor, Spink3.
Attitudes about psychotherapy: A qualitative study of introductory psychology students who have never been in psychotherapy and the influence of attachment style
Authors: Hill Clara E. , Satterwhite Dorli B. , Larrimore Maria L. , Mann Aliya R. , Johnson Victoria C. , Simon Rachel E. , Simpson Alexandra C. , Knox Sarah
Published: November 2011

Design: Interviews about attitudes about psychotherapy with 12 undergraduate students who had never been in psychotherapy were analysed using consensual qualitative research. Results: Participants believed that the client role is to disclose, be receptive, and be motivated; that the therapist role is to listen, support, and give advice; and that the therapeutic relationship should be close and personal. Participants had ideas about the benefits (a healing therapeutic relationship, personal and interpersonal changes) and the barriers (self‐stigma and public stigma, difficulty revealing, need to solve problems on own, cost) associated with seeking therapy, and they disliked the idea of being diagnosed. In contrast with participants who were securely attached, those who were insecurely attached more often wanted a professional therapeutic relationship, wanted the therapist to ask questions, mentioned fewer benefits to therapy, and thought that they would have difficulty disclosing to a therapist. Discussion: Implications for changing attitudes about psychotherapy and improving training programs for practitioners are discussed.
Study of mechanical characteristics of the knee extensor and flexor musculature of volleyball players
Authors: Rodríguez‐Ruiz David , Rodríguez‐Matoso Dario , Quiroga Miriam E. , Sarmiento Samuel , García‐Manso Juan Manuel , Da Silva‐Grigoletto Marzo E.
Published: November 2011

The aim of the present study was to analyse differences in muscle response and mechanical characteristics of the vastus medialis, rectus femoris, vastus lateralis, and biceps femoris in elite volleyball players of both sexes using tensiomyography. To this end, 47 players of nine nationalities playing for teams in the men's and women's Spanish Superleagues were assessed. The sample comprised 22 women (age 24.6±4.3 years; weight 72.14±10.06 kg; height 178.40±8.50 cm) and 25 men (age 25.0±4.3 years; weight 88.76±9.07 kg; height 194.71±7.84 cm). Tensiomyography was used to assess muscular response and muscular mechanical characteristics. For this purpose, the following variables were analysed: maximum radial displacement of muscle belly and normalized response speed. The findings show, both in men and women, a higher normalized response speed score in the vastus lateralis and vastus medialis compared with the rectus femoris and biceps femoris. A marked lateral symmetry of maximum radial displacement of the muscle belly was also observed in the musculature of the lower limbs, with no statistically significant differences being detected in either men or women. There were, however, clear differences in terms of normalized response speed between male and female volleyball players: women displayed a more pronounced difference in the normalized response speed of the musculature responsible for extension (vastus medialis, rectus femoris, and vastus lateralis) and flexion (biceps femoris) of the knee joint than men. Moreover, tensiomyography proved to be a highly sensitive tool for detecting such changes.
Water immersion as a recovery aid from intermittent shuttle running exercise
Authors: Corbett Jo , Barwood Martin J. , Lunt Heather C. , Milner Andrew , Tipton Michael J.
Published: November 2011

Aim: To investigate the effect of different water immersion (WI) treatments on recovery from intermittent shuttle running exercise in comparison to an ecologically relevant control.

Methods: Forty males performed 90 minutes intermittent shuttle running, following which they were assigned to either: (1) 12‐min standing WI at 12°C; (2) 12‐min standing WI at 35°C; (3) 2‐min seated WI at 12°C; (4) an ecologically relevant control consisting of 12 minutes walking at 5 km h−1. Muscle soreness, maximal voluntary contraction (MVC) of the knee flexors and extensors, hop distance, creatine kinase activity and myoglobin concentration were measured before exercise and in the 168 hours following the intervention. Between‐group differences, time effects and interaction effects were investigated by mixed‐model ANOVA.

Results: The shuttle running exercise induced an increase in muscle soreness (1, 24, 48 and 72 hours post‐intervention) creatine kinase activity and myoglobin concentration (post‐exercise and 1, 24 and 48 hours post‐intervention), and reduced MVC of the knee extensors (11% reduction at 24 hours, remaining reduced at 48 and 72 hours), flexors (24% reduction at 24 hours, remaining reduced at 24, 48, 72 and 168 hours), and hop distance (24 and 48 hours). However, no between‐group differences or interaction effects were evident for any of these parameters.

Conclusion: The WI protocols investigated were not better than light exercise in facilitating recovery from shuttle running exercise. Future studies examining the efficacy of WI as a recovery intervention should include a representative control condition to increase their relevance to sporting populations.
Team selection after a short cricket series
Authors: Lemmer Hermanus Hofmeyr
Published: November 2011

The selection of a cricket team cannot be fair unless the best available performance measures are used. The traditional batting average can be very unrealistic, especially in the case of a small number of scores with a high proportion of not out scores. In the present study the focus is on using the most suitable measures for the selection of a team after a small number of matches had been played. Provision is made for the fact that match conditions may influence the scoring rate of batsmen. These measures are used for illustration purposes to select a team from the players who played in the International Cricket Council (ICC) Champions Trophy 2009 One‐Day International (ODI) Series. It is shown how an integer programming method can be used for the selection process. The approach is that a well balanced cricket team should include different kinds of specialists, namely batsmen, bowlers, all‐rounders and a wicket‐keeper. A selection committee may be able to rank batsmen in order of batting ability and bowlers according to bowling ability, but when it comes to all‐rounders it is not so simple. The fact that an all‐rounder is, by definition, a good batsman and also a good bowler, makes it difficult to rank all‐rounders. Furthermore, how many of each specialist type should be selected? The purpose of this paper is to show how integer optimisation, an objective scientific method, can be used to aid in selecting a cricket team. Guidelines are also given for the selection of a team if career performance data have to be used.
Percentile values for flexibility tests in youths aged 6 to 17 years: Influence of weight status
Authors: Castro‐Piñero José , Girela‐Rejón María José , González‐Montesinos José Luis , Mora Jesús , Conde‐Caveda Julio , Sjöström Michael , Ruiz Jonatan R.
Published: November 2011

The aim of the present study was to examine the influence of weight status on flexibility performance in Spanish youths (1475 boys and 1237 girls) aged 6–17 years, and to provide percentile values for two flexibility tests. Flexibility was assessed by the sit‐and‐reach and shoulder stretch tests. Participants were categorized as underweight, normal weight, overweight and obese. Girls had significantly better scores than boys in all the studied tests, except in the right‐shoulder stretch test in the 6–7, 14–15 and 16–17‐year‐old groups, and in the left‐shoulder stretch test in the 6–7, 8–9 and 16–17‐year‐old groups. There was not a significant positive association between weight status and flexibility assessed by the sit‐and‐reach test. However, overweight and obese youths had significantly lower performance than their normal‐weight counterparts in the shoulder stretch test. Underweight and normal‐weight individuals had similar performances for both shoulder flexibility tests. In conclusion, percentiles values of the studied flexibility tests might be of interest to estimate the proportion of youths with high or low flexibility levels, as well as to evaluate the effects of alternative interventions.
No effect of upper body compression garments in elite flat‐water kayakers
Authors: Dascombe Ben , Laursen Paul , Nosaka Kazunori , Polglaze Ted
Published: November 2011

While the effect of lower body compression garments on performance and physiological responses are well documented, no studies have examined the effect of upper body compression garments (UBCG) on upper‐body dominant exercise. This study examined the effects of wearing UBCG on performance and physiological responses during simulated flat‐water kayaking. Five male (mean values±s: 21.8±2.8 years; 83.5±9.2 kg; 63.0±5.5 ml·kg−1·min−1) and two female (mean values±s: 25.0±4.2 years; 71.4±2.7 kg; 51.0±4.8 ml·kg‐1·min‐1) elite flat‐water kayakers completed a six‐step incremental test followed by a four‐minute maximal performance test (4minPT) in both UBCG and control (no shirt or sports training bra) conditions in a randomized counter‐balanced order. Heart rate and oxygen consumption (O2) as well as performance measures (power, distance covered, stroke rate) were recorded during the tests, and blood lactate was measured immediately after each incremental step and three minutes following the 4minPT. Near‐infrared spectroscopy‐derived measures of blood flow and oxygenation of the flexor carpi radialis were monitored continuously for all tests. No significant differences between the UBCG and control conditions were evident for any performance, cardiorespiratory or oxygenation measure across the incremental step test and 4minPT. It was concluded that wearing UBCG did not provide any significant physiological or performance benefits during simulated flat‐water kayaking.
Precision of the optimized carbon monoxide rebreathing method to determine total haemoglobin mass and blood volume
Authors: Durussel Jérôme , Ross Ramzy , Kodi Prithvi Raj , Daskalaki Evangelia , Takas Pantazis , Wilson John , Kayser Bengt , Pitsiladis Yannis
Published: November 2011

The aim of this study was to evaluate the influence of different apparatuses, procedures and calculations on the precision of the optimized carbon monoxide method. Total haemoglobin mass was determined twice on consecutive days in 11 subjects using both venous and capillary blood samples. To estimate loss of carbon monoxide due to exhalation, carbon monoxide concentration was measured by two portable carbon monoxide analysers (Fluke CO‐220, Fluke, Norwich, UK and Pac 7000 Carbon Monoxide, Draeger Safety, Northumberland, UK) and alveolar ventilation was specified using an automated metabolic gas analysis system (Cosmed Quark b2, Cosmed, Rome, Italy). Blood volume was derived from total haemoglobin mass using haemoglobin concentration and haematocrit obtained in both the supine and the seated position. Two different formulae to calculate blood volume were also compared. Precision was good for both total haemoglobin mass and blood volume measurements performed on consecutive days (typical error < 2%). Using Fluke CO‐220 analyser, an estimated alveolar ventilation and capillary blood, total haemoglobin mass (917±136 g) was similar when compared to the Pac 7000 Draeger CO‐analyser (904±137 g; mean bias –13 g with 95% limits of agreement –26 to + 1 g, P=0.76), specified alveolar ventilation (911±132 g, mean bias –6 g with 95% limits of agreement –18 g to + 6 g, P =0.87) and venous blood (917±134 g, mean bias 0 g with 95% limits of agreement –38 to + 38 g, P=0.99), respectively. Blood volume determination was also not significantly affected by the supine vs. seated position (6.7±0.8 l vs. 6.6±0.8 l, P=0.56) but can deviate by ~0.6 l (P=0.01) depending on the formula applied. Thus, the good precision of the assessment of total haemoglobin mass and blood volume using the optimized carbon monoxide rebreathing method is not significantly influenced by the make of CO analyser, method of obtaining alveolar ventilation, blood sampling method and subject position, but should for longitudinal monitoring purposes use the same formula.
Influence of electrically assisted cycling on physiological parameters in untrained subjects
Authors: de Geus Bas , Kempenaers Farid , Lataire Philippe , Meeusen Romain
Published: November 2011

Electrically assisted bicycles (EAB) as a form of transport not only offer the potential to reduce energy use and environmental impact but could also be an effective way of encouraging active living. The purpose of this study is to assess the influence of physical active commuting to work using an EAB, on physiological parameters in 20 untrained men and women. Tests were performed at three different time points over a 10‐week period, including four weeks of passive (control period) and six weeks of active commuting (intervention period). ANOVA for repeated measures was used to test differences between the testing series for the most important physiological parameters: Pmax·kg−1, V˙O2peak·kg−1, fixed blood lactate concentration (2, 4 mmol·l−1). The subjects performed over a 6‐week period a mean total cycling distance of 405.1±156.0 km with a weekly frequency of 4.1±1.7 days·week−1 for men and 246.0±116.3 km with a frequency of 2.9±1.0 days·week−1 for women. Pmax·kg−1 significantly increased in men and women after 6 weeks of active commuting. Power output at 2 mmol·l−1 significantly increased in women (P=0.001) but not in men (P=0.0604). Power output at 4 mmol·l−1 significantly increased for men and women. V˙O2peak·kg−1 did not differ. With respect to the study limitations, it is concluded that cycling to work on an EAB was effective in increasing the maximal power output and power output at 4 mmol·l−1 in these untrained subjects. Cycling on an EAB seems to be a promising tool as a health enhancing physical activity, for those who will benefit the most in terms of health related fitness, namely the physically inactive, unfit and older people.
Ranking benchmarks of top 100 players in men's professional tennis
Authors: Reid Machar , Morris Craig
Published: November 2011

In men's professional tennis, players aspire to hold the top ranking position. On the way to the top spot, reaching the top 100 can be seen as a significant career milestone. National Federations undertake extensive efforts to assist their players to reach the top 100. However, objective data considering reasonable ranking yardsticks for top 100 success in men's professional tennis are lacking. Therefore, it is difficult for National Federations and those involved in player development to give empirical programming advice to young players. By taking a closer look at the ranking history of professional male tennis players, this article tries to provide those involved in player development a more objective basis for decision‐making. The 100 names, countries, birthdates and ranking histories of the top 100 players listed in the Association of Tennis Professionals (ATP) at 31 December 2009 were recorded from websites in the public domain. Descriptive statistics were reported for the ranking milestones of interest. Results confirmed the merits of the International Tennis Federation's junior tour with 91% of the top 100 professionals earning a junior ranking, the mean peak of which was 94.1, s=148.9. On average, top 100 professionals achieved their best junior rankings and earned their first ATP point at similar ages, suggesting that players compete on both the junior and professional tours during their transition. Once professionally ranked, players took an average 4.5, s=2.1 years to reach the ATP top 100 at the mean age of 21.5, s=2.6 years, which contrasts with the mean current age of the top 100 of 26.8, s=3.2. The best professional rankings of players born in 1982 or earlier were positively related to the ages at which players earned their first ATP point and then entered the top 100, suggesting that the ages associated with these ranking milestones may have some forecasting potential. Future work should focus on the change in top 100 demographics over time as well as the evaluation of the interaction between rankings and tournament play.
Follicular dendritic cells help establish follicle identity and promote B cell retention in germinal centers
Authors: Wang Xiaoming , Cho Bryan , Suzuki Kazuhiro , Xu Ying , Green Jesse A. , An Jinping , Cyster Jason G.
Published: November 2011

Follicular dendritic cells (FDCs) retain and display opsonized antigens in primary follicles and germinal centers (GCs). However, their roles beyond antigen presentation have been incompletely defined. In this study, we tested the impact of selective FDC ablation on short-term follicle and GC function. Within 2 d of FDC ablation, primary follicles lost their homogeneity and became disorganized bands of cells around T zones. These B cell areas retained CXCL13-expressing stromal cells but often exhibited inappropriate ER-TR7 and CCL21 expression. Ablation of GC FDCs led to the disappearance of GCs. When B cell death was prevented using a Bcl2 transgene, FDC ablation led to splenic GC B cell dispersal. Mesenteric lymph node GCs were more resistant but became dispersed when sphingosine-1-phosphate receptor-2 was also removed. These experiments indicate that FDCs help maintain primary follicles as a B cell exclusive niche and define a critical role for FDCs in cell retention within GCs.
The exceptional set in Vojta’s conjecture for algebraic points of bounded degree
Authors: Levin Aaron
Published: November 2011

We study the dependence on various parameters of the exceptional set in Vojta’s conjecture. In particular, by making use of certain elliptic surfaces, we answer in the negative the often-raised question of whether Vojta’s conjecture holds when extended to all algebraic points (that is, if the conjecture holds without fixing a bound on the degree of the algebraic points).
A Review of Huanglongbing (Citrus Greening) Management in Citrus Using Nutritional Approaches in China
Authors: Xia Yulu , Ouyang Gecheng , Sequeira Ronald A. , Takeuchi Yu , Baez Ignacio , Chen Jianchi
Published: November 2011

The Asian form of huanglongbing (HLB) is caused by ‘Candidatus Liberibacter asiaticus (Las),’ a phloem-limited bacterium transmitted by the Asian citrus psyllid, Diaphorina citri Kuwayama. Nutrient management, together with other cultural practices such as pruning and irrigation, for mitigation of the disease has been practiced in China for many years. Our literature review, field survey, and interviews with Chinese scientists and growers indicate that these cultural practices were generally ineffective for the disease management. However, a nutritional approach in conjunction with other cultural practices such as irrigation can maintain grove productivity for a certain time depending on the type of citrus species/cultivars, the age of the trees, the propagation method of the plants, the Asian citrus psyllid (ACP) (Diaphorina citri Kuwayama) population, and other factors. Symptomatic mature pommelo (Citrus maxima Merr) and sweet orange (C. sinensis L. Osbeck) plants can commonly survive and maintain a certain level of productivity for an additional 4 to 5 years, even longer assuming vigorous ACP control.

Accepted for publication 27 June 2011. Published 3 October 2011.
Biochemical and Ultrastructural Changes in the Hepatopancreas ofBellamya aeruginosa(Gastropoda) Fed with Toxic Cyanobacteria
Authors: Zhu Jinyong , Lu Kaihong , Zhang Chunjing , Liang Jingjing , Hu Zhiyong
Published: November 2011

This study was conducted to investigate ultrastructural alterations and biochemical responses in the hepatopancreas of the freshwater snailBellamya aeruginosaafter exposure to two treatments: toxic cyanobacterium (Microcystis aeruginosa) and toxic cyanobacterial cells mixed with a non-toxic green alga (Scendesmus quadricauda) for a period of 15 days of intoxication, followed by a 15-day detoxification period. The toxic algal suspension induced a very pronounced increase of the activities of acid phosphatases, alkaline phosphatases and glutathione S-transferases (ACP, ALP and GST) in the liver at the later stage of intoxication. During the depuration, enzymatic activity tended to return to the levels close to those in the control. The activity of GST displayed the most pronounced response among different algal suspensions. Severe cytoplasmic vacuolization, condensation and deformation of nucleus, dilation and myeloid-like in mitochondria, disruption of rough endoplasmic reticulum, proliferation of lysosome, telolysosomes and apoptotic body were observed in the tissues. All cellular organelles began recovery after the snails were transferred to theS. quadricauda.The occurrence of a large amount of activated lysosomes and heterolysosomes and augment in activity of detoxification enzyme GST might be an adaptive mechanism to eliminate or lessen cell damage caused by hepatotoxicity toB. aeruginosa.
Nonlinear Model-Based Method for Clustering Periodically Expressed Genes
Authors: Tian Li-Ping , Liu Li-Zhi , Zhang Qian-Wei , Wu Fang-Xiang
Published: November 2011

Clustering periodically expressed genes from their time-course expression data could help understand the molecular mechanism of those biological processes. In this paper, we propose a nonlinear model-based clustering method for periodically expressed gene profiles. As periodically expressed genes are associated with periodic biological processes, the proposed method naturally assumes that a periodically expressed gene dataset is generated by a number of periodical processes. Each periodical process is modelled by a linear combination of trigonometric sine and cosine functions in time plus a Gaussian noise term. A two stage method is proposed to estimate the model parameter, and a relocation-iteration algorithm is employed to assign each gene to an appropriate cluster. A bootstrapping method and an average adjusted Rand index (AARI) are employed to measure the quality of clustering. One synthetic dataset and two biological datasets were employed to evaluate the performance of the proposed method. The results show that our method allows the better quality clustering than other clustering methods (e.g.,k-means) for periodically expressed gene data, and thus it is an effective cluster analysis method for periodically expressed gene data.
Overlapping, Additive and Counterregulatory Effects of Type II and I Interferons on Myeloid Dendritic Cell Functions
Authors: Frasca Loredana , Lande Roberto
Published: November 2011

Dendritic cells (DCs) are central player in immunity by bridging the innate and adaptive arms of the immune system (IS). Interferons (IFNs) are one of the most important factors that regulate both innate and adaptive immunity too. Thus, the understanding of how type II and I IFNs modulate the immune-regulatory properties of DCs is a central issue in immunology. In this paper, we will address this point in the light of the most recent literature, also highlighting the controversial data reported in the field. According to the wide literature available, type II as well as type I IFNs appear, at the same time, to collaborate, to induce additive effects or overlapping functions, as well as to counterregulate each one's effects on DC biology and, in general, the immune response. The knowledge of these effects has important therapeutic implications in the treatment of infectious/autoimmune diseases and cancer and indicates strategies for using IFNs as vaccine adjuvants and in DC-based immune therapeutic approaches.
Average group behavior does not represent individual behavior in classical conditioning of the honeybee
Authors: Pamir Evren , Chakroborty Neloy Kumar , Stollhoff Nicola , Gehring Katrin Barbara , Antemann Victoria , Morgenstern Laura , Felsenberg Johannes , Eisenhardt Dorothea , Menzel Randolf , Nawrot Martin Paul
Published: November 2011

Conditioned behavior as observed during classical conditioning in a group of identically treated animals provides insights into the physiological process of learning and memory formation. However, several studies in vertebrates found a remarkable difference between the group-average behavioral performance and the behavioral characteristics of individual animals. Here, we analyzed a large number of data (1640 animals) on olfactory conditioning in the honeybee (Apis mellifera). The data acquired during absolute and differential classical conditioning differed with respect to the number of conditioning trials, the conditioned odors, the intertrial intervals, and the time of retention tests. We further investigated data in which animals were tested for spontaneous recovery from extinction. In all data sets we found that the gradually increasing group-average learning curve did not adequately represent the behavior of individual animals. Individual behavior was characterized by a rapid and stable acquisition of the conditioned response (CR), as well as by a rapid and stable cessation of the CR following unrewarded stimuli. In addition, we present and evaluate different model hypotheses on how honeybees form associations during classical conditioning by implementing a gradual learning process on the one hand and an all-or-none learning process on the other hand. In summary, our findings advise that individual behavior should be recognized as a meaningful predictor for the internal state of a honeybee—irrespective of the group-average behavioral performance.
Preparation of Cuticles from Feeding Drosophila Larvae: Figure 1.
Authors: Stern David L. , Sucena Elio
Published: November 2011

The Drosophila cuticle carries a rich array of morphological details. Thus, cuticle examination has had a central role in the history of genetics. Studies of the Drosophila cuticle have focused mainly on first-instar larvae and adult cuticular morphology. Although the cuticles of second- and third-instar larvae are strikingly different from those of the first instar, these differences have been poorly studied. This protocol describes three methods for preparing cuticles from fed larvae. One commonly used procedure involves manually pricking the larvae. A simpler method for preparing larval cuticles is to burst the larvae once they have been mounted. This method is used for first- and second-instar larvae and does not require pricking; it removes the gut contents by “popping” the rear of the embryo using pressure from the coverslip. If just the right amount of medium is used, the coverslip will be pulled toward the slide, applying pressure on the samples. The larvae usually burst from their posterior ends. Also presented is an alternative procedure designed specifically for the use with third-instar larvae, although the “pricking” method can be used at this stage.
Preparation of Drosophila Eye Specimens for Scanning Electron Microscopy
Authors: Wolff Tanya
Published: November 2011

Scanning electron microscopy (SEM) allows for high-magnification analysis of the external structures of the Drosophila eye. In this article, three methods are provided: one for eyes that are to be critical-point-dried (CPD) and two alternatives if a critical point drier is not available. The major difference between the first method and the other two is a preliminary fixation step. Drosophila eyes have a tendency to collapse when CPD, so fixing the animals before critical point drying improves the yield of good specimens.
Conditioning to Colors: A Population Assay for Visual Learning in Drosophila: Figure 1.
Authors: van Swinderen Bruno
Published: November 2011

Vision is a major sensory modality in Drosophila behavior, with more than one-half of the Drosophila brain devoted to visual processing. The mechanisms of vision in Drosophila can be studied in individuals and in populations of flies by using various paradigms. Although there has never been a widely used population assay for visual learning in Drosophila, some population paradigms have shown significant visual learning. These studies use colors as conditioned stimuli (CS) and shaking as the unconditioned stimulus (US). A simple version of the paradigm, conditioning to colors using a shaking device, is described here. A conditioning chamber, called a crab, is designed to center the flies after shaking by having them tumble down to the lowest point between joined glass tubes forming a V. Thus, vibration should be just strong enough to center most flies. After shaking, flies display a geotactic response and climb up either side of the V, and their choice of which side to climb is influenced by color displays on either side. The proportion of flies on either side determines the flies’ natural preference or their learned avoidance of a color associated with shaking.
The Optomotor Maze: A Population Assay for Visual Perception in Drosophila: Figure 1.
Authors: van Swinderen Bruno
Published: November 2011

Vision is a major sensory modality in Drosophila behavior, with more than one-half of the Drosophila brain devoted to visual processing. The mechanisms of vision in Drosophila can now be studied in individuals and in populations of flies by using various paradigms. The optomotor maze, described here, is a novel and efficient approach for querying visual perception in Drosophila populations. The optomotor maze setup is very simple: An eight-choice maze consisting of 3-mm paths grooved into a transparent Plexiglas or acrylic slab is placed over an upturned cathode ray tube (CRT) monitor on which visuals are displayed. The placement or movement of the visuals on the CRT, which the flies can see through the flat bottom of the maze, influences their turning behavior at each choice point. This paradigm can be adapted for visual learning by simply rerunning flies in the maze (habituation) or as a more sophisticated version of the aversive phototaxic suppression (APS) paradigm.
Photobleaching Regions of Living Cells to Monitor Membrane Traffic
Authors: Snapp Erik Lee , Lajoie Patrick
Published: November 2011

Eukaryotic cells are composed of an intricate system of internal membranes that are organized into different compartments—including the endoplasmic reticulum (ER), the nuclear envelope, the Golgi complex (GC), lysosomes, endosomes, caveolae, mitochondria, and peroxisomes—that perform specialized tasks within the cell. The localization and dynamics of intracellular compartments are now being studied in living cells because of the availability of green fluorescent protein (GFP)-fusion proteins and recent advances in fluorescent microscope imaging systems. This protocol outlines two methods for photobleaching living cells to monitor membrane traffic. The first method involves selective photobleaching using a confocal laser-scanning microscope (CLSM) that can bleach discrete selected regions of interest. As outlined in the second method, photobleaching can also be performed with older CLSMs that lack the capacity for selective photobleaching. In this case, photobleaching is accomplished by zooming into a small region of the cell and scanning with full laser power.
Time-Lapse Imaging of Fluorescently Labeled Live Cells in the Embryonic Mammalian Forebrain
Authors: Noctor Stephen C.
Published: November 2011

Time-lapse imaging of fluorescently labeled cells in organotypic slice culture provides a unique window through which to view development of the embryonic forebrain. The live imaging approach allows investigators to directly observe and record neurogenic and gliogenic divisions in the developing forebrain, to record the proliferative behavior of embryonic precursor cells, and to track newborn cells as they migrate from the proliferative zones to their final destination. This approach allows for identification and characterization of embryonic precursor cells, permits the physiological and immunohistochemical characterization of the fluorescent cells under study, and presents an opportunity to test hypotheses about mechanisms that guide developmental processes in the forebrain. This article describes a protocol for time-lapse imaging of fluorescently labeled cells in the embryonic forebrain of organotypic slice cultures. The procedure does not require prohibitively expensive on-stage incubation systems, and can yield reliable data from embryonic slices for up to 1 wk. Tracking the movement of labeled cells in slice culture is relatively straightforward in those cases in which there are few labeled cells. Although this approach is time intensive, it provides beautifully detailed images that reveal morphological and functional data from labeled cells over time, and can yield a significant amount of data from each cell.
Preparation and 4D Fluorescent Imaging of Quail Embryos
Authors: Canaria Christie A. , Lansford Rusty
Published: November 2011

Traditionally, our understanding of developmental biology has been based on the fixation and study of embryonic samples. Detailed microscopic scrutiny of static specimens at varying ages allowed for anatomical assessment of tissue development. The advent of confocal and two-photon excitation (2PE) microscopy enables researchers to acquire volumetric images in three dimensions (x, y, and z) plus time (t). The development of transgenic Japanese quail has provided an embryonic model system that is more easily accessible than mammalian models and more efficient to breed than the classic avian model, the chicken. This protocol describes the methods necessary to prepare transgenic quail embryos for imaging in vitro, and for acquiring, processing, and analyzing three-dimensional (3D) time-lapse images.
In Vivo Transfection Using Cyclodextrin-Containing Polycations
Authors: Heidel Jeremy D.
Published: November 2011

Numerous nonviral systems have been developed for the delivery of nucleic acids to cultured cells and to particular cell types in vivo. These systems vary with regard to their toxicity, immunogenicity, and ability to target particular cell surface receptors and/or cell types. A class of linear cationic polymers containing the sugar β-cyclodextrin (β-CD) has been shown to be effective at delivering a variety of nucleic acids in vivo, including plasmid DNA, DNAzymes, and short interfering RNAs (siRNAs). These polymer–nucleic acid complexes (polyplexes) can be further modified to incorporate a targeting ligand such as transferrin to induce preferential uptake of polyplexes by cells expressing high levels of the cognate receptor. This protocol describes a procedure for the use of cyclodextrin-containing polycations (CDPs) in vivo. Salt stabilization and cell targeting are critical to the success of in vivo transfection using CDPs, so adamantane-poly(ethylene glycol) (AD-PEG) conjugates (both unmodified AD-PEG and an AD-PEG-Ligand conjugate) are included in these formulations. The amount of the AD-PEG-Ligand conjugate included depends on numerous factors, including its effect on polyplex stability (influenced by ligand size and charge) and the density of the cognate receptor on target cell type(s). Some targeting ligands may have extreme sizes or net charges that could present a challenge to their incorporation into these polyplex formulations.
Attachment of Anti-GFP Antibodies to Microspheres for Optical Trapping Experiments
Authors: Spudich James A. , Rice Sarah E. , Rock Ronald S. , Purcell Thomas J. , Warrick Hans M.
Published: November 2011

In vitro motility assays enabled the analysis of coupling between ATP hydrolysis and movement of myosin along actin filaments or kinesin along microtubules. Single-molecule assays using laser trapping have been used to obtain more detailed information about kinesins, myosins, and processive DNA enzymes. The combination of in vitro motility assays with laser-trap measurements has revealed detailed dynamic structural changes associated with the ATPase cycle. This protocol describes a method for attaching anti-GFP (green fluorescent protein) antibodies to microspheres. GFP-motor fusion proteins can then be adsorbed to the microspheres for use in single-molecule motility studies and optical trapping experiments.
4D Fluorescent Imaging of Embryonic Quail Development: Table 1.
Authors: Canaria Christie A. , Lansford Rusty
Published: November 2011

Traditionally, our understanding of developmental biology has been based on the fixation and study of embryonic samples. Detailed microscopic scrutiny of static specimens at varying ages allowed for anatomical assessment of tissue development. The advent of confocal and two-photon excitation (2PE) microscopy enables researchers to acquire volumetric images in three dimensions (x, y, and z) plus time (t). Here, we present techniques for acquisition and analysis of three-dimensional (3D) time-lapse data. Both confocal microscopy and 2PE microscopy techniques are used. Data processing for tiled image stitching and time-lapse analysis is also discussed. The development of a transgenic Japanese quail system, as discussed here, has provided an embryonic model that is more easily accessible than mammalian models and more efficient to breed than the classic avian model, the chicken.
Cyclodextrin-Containing Polycations for Nucleic Acid Delivery
Authors: Heidel Jeremy D.
Published: November 2011

Numerous nonviral systems have been developed for the delivery of nucleic acids to cultured cells and to particular cell types in vivo. These systems vary with regard to their toxicity, immunogenicity, and ability to target particular cell surface receptors and/or cell types. A class of linear cationic polymers containing the sugar β-cyclodextrin has been shown to be effective at delivering a variety of nucleic acids in vivo, including plasmid DNA, DNAzymes, and short interfering RNAs (siRNAs). These polymer–nucleic acid complexes (polyplexes) can be further modified to incorporate a targeting ligand such as transferrin to induce preferential uptake of polyplexes by cells expressing high levels of the cognate receptor. This article presents background information on cyclodextrin-containing polycations.
Optical Traps to Study Properties of Molecular Motors
Authors: Spudich James A. , Rice Sarah E. , Rock Ronald S. , Purcell Thomas J. , Warrick Hans M.
Published: November 2011

In vitro motility assays enabled the analysis of coupling between ATP hydrolysis and movement of myosin along actin filaments or kinesin along microtubules. Single-molecule assays using laser trapping have been used to obtain more detailed information about kinesins, myosins, and processive DNA enzymes. The combination of in vitro motility assays with laser-trap measurements has revealed detailed dynamic structural changes associated with the ATPase cycle. This article describes the use of optical traps to study processive and nonprocessive molecular motor proteins, focusing on the design of the instrument and the assays to characterize motility.
Isolation and molecular characterization of methicillin‐resistant Staphylococcus aureus from public transport
Authors: Iwao Yasuhisa , Yabe Shizuka , Takano Tomomi , Higuchi Wataru , Nishiyama Akihito , Yamamoto Tatsuo
Published: November 2011

Methicillin‐resistant Staphylococcus aureus (MRSA) not only causes disease in hospitals, but also in the community. The characteristics of MRSA transmission in the environment remain uncertain. In this study, MRSA were isolated from public transport in Tokyo and Niigata, Japan. Of 349 trains examined, eight (2.3%) were positive for MRSA. The MRSA isolated belonged to sequence types (STs) 5, 8, 88, and 89, and included community infection‐associated ST8 MRSA (with novel type IV staphylococcal cassette chromosome mec) and the ST5 New York/Japan hospital clone. The data indicate that public transport could contribute to the spread of community‐acquired MRSA, and awareness of this mode of transmission is necessary.
Helicobacter pylori antigens as potential modulators of lymphocytes’ cytotoxic activity
Authors: Rudnicka Karolina , Włodarczyk Marcin , Moran Anthony P. , Rechciński Tomasz , Miszczyk Eliza , Matusiak Agnieszka , Szczęsna Ewelina , Walencka Maria , Rudnicka Wiesława , Chmiela Magdalena
Published: November 2011

Helicobacter pylori (H.p) colonizes human gastric mucosa and causes gastric and duodenal ulcer disease or gastric cancer. Various H.p compounds may modulate the host immune response in regards to tolerance of the infection or disease development. The aim of this study was to determine whether H.p lipopolysaccharide (LPS) and glycine acid extract antigens (GE) or E. coli LPS influence the cytotoxic activity of peripheral blood lymphocytes from H.p infected – H.p (+) or uninfected – H.p (−) individuals, in the presence or absence of exogenous interleukin (IL)12. Individual H.p status was defined by the urea breath test. Lymphocytes, stimulated or not with H.p, and control antigens, with or without IL‐12, were used as effector cells and epithelial HeLa cells as targets. The cytotoxicity of lymphocytes was expressed as the percentage of dead target cells unable to reduce tetrazolium salt. The supernatants from HeLa/lymphocyte cultures were used for detection of the cellular cytotoxicity markers granzyme B and caspase 8. The natural cytotoxic activity of lymphocytes from H.p (+) was less than that of H.p (−) donors. This may have been due to fewer natural killer cells of CD3−CD56+Nkp46+ phenotype in H.p (+) in comparison to H.p (−) subjects. H.p GE and standard E. coli LPS enhanced the cytotoxicity of lymphocytes towards target cells whereas H.p LPS downregulated this activity. The decrease in lymphocyte cytotoxicity in response to H.p LPS correlated with a lack of IL‐2 and IL‐12 production, inhibition of interferon‐γ production, and low IL‐10 secretion by mononuclear leukocytes. IL‐12 significantly enhanced the natural as well as H.p LPS and H.p GE driven cytotoxic capacity of lymphocytes. In conclusion, H.p LPS may negatively modulate natural cytotoxic activity and cytokine secretion by immunocompetent cells and thus be involved in the maintenance of infection and development of gastric pathologies.
The association between maternal serious psychological distress and child obesity at 3 years: a cross‐sectional analysis of the UK Millennium Cohort Data
Authors: Ramasubramanian L. , Lane S. , Rahman A.
Published: November 2011

Background  The prevalence of child obesity is increasing rapidly worldwide. Early childhood has been identified as a critical time period for the development of obesity. Maternal mental health and early life environment are crucial factors and have been linked to adverse child outcomes. The objective of the study was to examine the relationship between maternal serious psychological distress and obesity in early childhood.

Methods  A cross‐sectional analysis of data from the Millennium Cohort Study was conducted. Subjects consisted of all natural mothers (n= 10 465) who had complete and plausible data for Kessler‐6 scores, socio‐demographic and anthropometric variables, and their children for whom anthropometric measurements were completed at age 3. Maternal serious psychological distress was defined as a score of 13 or more on the Kessler‐6 scale. Obesity was defined as body mass index ≥95th centile of the 1990 reference chart for age and sex in children. The data were analysed using spss 16. Maternal socio‐demographic factors that are known to influence maternal mental health and child obesity were identified and adjusted using multivariate logistic regression.

Results  Of the 10 465 mother–child dyads, 3.5% of mothers had serious psychological distress and 5.5% of children were obese at 3 years of age. Logistic regression analysis showed that maternal serious psychological distress was associated with early childhood obesity (P= 0.01; OR 1.62, 95% CI 1.11, 2.37). After adjusting for potential confounding factors using multivariate logistic regression, maternal serious psychological distress remained significantly associated with early childhood obesity (P= 0.01; OR 1.59, 95% CI 1.08, 2.34).

Conclusions  The results show that maternal serious psychological distress is independently associated with early childhood obesity.