Internal and external validity of cluster randomised trials: systematic review of recent trials

Objectives To assess aspects of the internal validity of recently published cluster randomised trials and explore the reporting of information useful in assessing the external validity of these trials.Design Review of 34 cluster randomised trials in primary care published in 2004 and 2005 in seven journals (British Medical Journal, British Journal of General Practice, Family Practice, Preventive Medicine, Annals of Internal Medicine, Journal of General Internal Medicine, Pediatrics).Data sources National Library of Medicine (Medline) via PubMed.Data extraction To assess aspects of internal validity we extracted data on appropriateness of sample size calculations and analyses, methods of identifying and recruiting individual participants, and blinding. To explore reporting of information useful in assessing external validity we extracted data on cluster eligibility, cluster inclusion and retention, cluster generalisability, and the feasibility and acceptability of the intervention to health providers in clusters.Results 21 (62%) trials accounted for clustering in sample size calculations and 30 (88%) in the analysis; about a quarter were potentially biased because of procedures surrounding recruitment and identification of patients; individual participants were blind to allocation status in 19 (56%) and outcome assessors were blind in 15 (44%). In almost half the reports, information relating to generalisability of clusters was poorly reported, and in two fifths there was no information about the feasibility and acceptability of the intervention.Conclusions Cluster randomised trials are essential for evaluating certain types of interventions. Issues affecting their internal validity, such as appropriate sample size calculations and analysis, have been widely disseminated and are now better addressed by researchers. Blinding of those identifying and recruiting patients to allocation status is recommended but is not always carried out. There may be fewer barriers to internal validity in trials in which individual participants are not recruited. External validity seems poorly addressed in many trials, yet is arguably as important as internal validity in judging quality as a basis for healthcare intervention.     

OsTDL1A binds to the LRR domain of rice receptor kinase MSP1, and is required to limit sporocyte numbers

Hybrids lose heterotic yield advantage when multiplied sexually via meiosis. A potential alternative breeding system for hybrids is apospory, where female gametes develop without meiosis. Common among grasses, apospory begins in the nucellus, where aposporous initials (AIs) appear near the sexual megaspore mother cell (MeMC). The cellular origin of AIs is obscure, but one possibility, suggested by the mac1 and msp1 mutants of maize and rice, is that AIs are apomeiotic derivatives of the additional MeMCs that appear when genetic control over sporocyte numbers is relaxed. MULTIPLE SPOROCYTES1 (MSP1) encodes a leucine-rich-repeat receptor kinase, which is orthologous to EXS/EMS1 in Arabidopsis. Like mac1 and msp1, exs/ems1 mutants produce extra sporocytes in the anther instead of a tapetum, causing male sterility. This phenotype is copied in mutants of TAPETUM DETERMINANT1 (TPD1), which encodes a small protein hypothesized to be an extracellular ligand of EXS/EMS1. Here we show that rice contains two TPD1-like genes, OsTDL1A and OsTDL1B. Both are co-expressed with MSP1 in anthers during meiosis, but only OsTDL1A and MSP1 are co-expressed in the ovule. OsTDL1A binds to the leucine-rich-repeat domain of MSP1 in yeast two-hybrid assays and bimolecular fluorescence complementation in onion cells; OsTDL1B lacks this capacity. When driven by the maize Ubiquitin1 promoter, RNA interference against OsTDL1A phenocopies msp1 in the ovule but not in the anther. Thus, RNAi produces multiple MeMCs without causing male sterility. We conclude that OsTDL1A binds MSP1 in order to limit sporocyte numbers. OsTDL1A-RNAi lines may be suitable starting points for achieving synthetic apospory in rice.     

Synthesis and structure-activity relationships of selective norepinephrine reuptake inhibitors (sNRI) with improved pharmaceutical characteristics

The design synthesis and SAR of a series of chiral ring-constrained norepinephrine reuptake inhibitors with improved physicochemical properties is described. Typical compounds are potent (IC(50)s<10 nM), selective against the other monoamine transporters, weak CYP2D6 inhibitors (IC(50)s>1 microM) and stable to oxidation by human liver microsomes. In addition, the compounds exhibit a favorable polarity profile.     

Synthesis and structure-activity relationships of selective norepinephrine reuptake inhibitors (sNRI) with a heterocyclic ring constraint

The design, synthesis and SAR of a series of heterocyclic ring-constrained norepinephrine reuptake inhibitors are described. As racemates, the best compounds compare favorably with atomoxetine (IC(50)'s<10 nM) in potency at the transporter.     

The conservation of the belted beauty Lycia zonaria britannica (Lepidoptera, Geometridae) in the United Kingdom

Recent surveys have established that the belted beauty moth Lycia zonaria britannica, which is an endemic subspecies found only in the British Isles, now occurs at just three sites in England and Wales. Populations at all three sites are small, with annual peak counts of usually less than 50 females. At Morfa Conwy in Wales and Meols Common in England, habitat quality has been declining for more than a decade as open, herb-rich, early successional dune grassland is replaced by fixed, rank grassland. Uniquely at Sunderland Point in England, the moth is primarily associated with areas of saltmarsh, but distribution is constrained by surrounding farmland and man-made structures. Attempts are now being made to restore areas of open grassland and to return the moth to its former distribution. Since the winter of 2000-2001, small plots of rank grassland on Meols Common have been excavated annually, with all vegetation removed, together with any humic layer present. The exposed bare sand has then been allowed to re-vegetate naturally. Within 18 months, the plots have developed into open, herb-rich habitats suitable for the moth. Adults were recorded on the plots for the first time in spring 2003 and may have been the product of the small numbers of eggs and larvae introduced to the plots in spring 2002. Similar excavations have been initiated at Morfa Conwy, which is in the process of being notified as a Site of Special Scientific Interest (SSSI). A 3-year programme to establish a second Welsh population is underway and, to date, more than 2000 captive-reared larvae have been released on the receptor site which lies 21 km to the east of Morfa Conwy.     

Sexual motivation suppresses paternal behaviour of male gerbils during their mates' postpartum oestrus

Adult male Mongolian gerbils, Meriones unguiculatus, avoid contact with their young on the day that the young are born. However, on succeeding days, fathers spend nearly as much time in contact with their offspring as do mothers. We undertook a series of studies to investigate the causes of the day-to-day change in male parental behaviour. In experiment 1, we tested males' response to pups before, after and on the day of their mates' parturition, and found that males were more parental both before and after the day of birth of their young than on that day. In experiment 2, we compared the parental behaviour of males paired either with intact or with ovariectomized dams (which do not come into postpartum oestrus) and found that males that were mated to intact females were less attentive to pups on the day of their birth than were the males mated to ovariectomized females. In experiment 3, we compared the parental responses of castrated and intact males to newborn pups and found that castrated males were more parental than intact males. In experiment 4, we compared the parental responses of males that were exposed to postpartum oestrous females but prevented from mating for 24 h. Extending the period of male sexual arousal to 24 h inhibited paternal responsiveness to neonates for 24 h. We interpret these results as being consistent with the hypothesis that on the day of birth of a litter, a male's parental behaviour is inhibited by the motivation to mate during his partner's postpartum oestrus.     

Upper and Lower Temperature Tolerances of Juvenile Freshwater Game-Fish Species Exposed to 32 days of Cycling Temperatures

Hydrobiologia - Temperature tolerance quantified as CTminima and CTmaxima of channel catfish (Ictalurus punctatus), largemouth bass (Micropterus salmoides) and rainbow trout (Oncorhynchus mykiss)...     

A biological assessment of Apophua simplicipes (Hymenoptera: Ichneumonidae) as a parasitoid of the oblique banded leafroller, Choristoneura rosaceana (Lepidoptera: Tortricidae)

Apophua simplicipes (Cresson) (Hymenoptera: Ichneumonidae) is a common parasitoid of the oblique banded leafroller, Choristoneura rosaceana (Harris) (Lepidoptera: Tortricidae) in organically managed apple orchards in the southern interior of British Columbia, Canada. The biological characteristics of a laboratory colony of A. simplicipes were studied. When held at 15 and 25°C and provided with honey water, individual females survived an average of 60.6±6.1 and 29.8±4.7 days, oviposited 196.7±50.7 and 326.6±51.3 eggs and parasitized a total of 163.4±40.4 and 229.4±35.8 hosts, respectively. Females oviposited into first through fourth instar oblique banded leafrollers, with significantly more parasitism occurring in the first two instars compared to the third and fourth instars. No parasitoid larvae survived past the first larval stage in parasitized fourth instar hosts. Apophua simplicipes did not parasitize larvae of three-lined leafroller, Pandemis limitata (Robinson) (Lepidoptera: Tortricidae) which is sympatric with oblique banded leafrollers in orchards in the southern interior of British Columbia. Female predation and host feeding from wounds on early instars of both leafroller species was observed under laboratory conditions. In addition, early instar hosts exited diet feeding sites in response to the probing activity of the ovipositing wasps. A similar escape reaction in the orchard may cause a leafroller larva to move away from its feeding site, making it more vulnerable to predation or movement off the tree. Apophua simplicipes larvae emerged from fifth and sixth instar hosts. Parasitized oblique banded leafroller hosts consumed significantly less meridic diet than unparasitized female larvae from fifth instar through to parasitoid emergence or leafroller pupation. Our laboratory results suggest that A. simplicipes may reduce field populations of oblique banded leafroller and decrease pest feeding damage.     

Antisense masking of an hnRNP A1/A2 intronic splicing silencer corrects SMN2 splicing in transgenic mice

Survival of motor neuron 2, centromeric (SMN2) is a gene that modifies the severity of spinal muscular atrophy (SMA), a motor-neuron disease that is the leading genetic cause of infant mortality. Increasing inclusion of SMN2 exon 7, which is predominantly skipped, holds promise to treat or possibly cure SMA; one practical strategy is the disruption of splicing silencers that impair exon 7 recognition. By using an antisense oligonucleotide (ASO)-tiling method, we systematically screened the proximal intronic regions flanking exon 7 and identified two intronic splicing silencers (ISSs): one in intron 6 and a recently described one in intron 7. We analyzed the intron 7 ISS by mutagenesis, coupled with splicing assays, RNA-affinity chromatography, and protein overexpression, and found two tandem hnRNP A1/A2 motifs within the ISS that are responsible for its inhibitory character. Mutations in these two motifs, or ASOs that block them, promote very efficient exon 7 inclusion. We screened 31 ASOs in this region and selected two optimal ones to test in human SMN2 transgenic mice. Both ASOs strongly increased hSMN2 exon 7 inclusion in the liver and kidney of the transgenic animals. Our results show that the high-resolution ASO-tiling approach can identify cis-elements that modulate splicing positively or negatively. Most importantly, our results highlight the therapeutic potential of some of these ASOs in the context of SMA.