bluuebunny/crossref_metadata_2025 huggingface dataset:
Effort, co‐operation and co‐ordination in an open source software project: GNOME
Authors: Koch Stefan , Schneider Georg
Published: March 2003

Abstract This paper presents results from research into open source projects from a software engineering perspective. The research methodology employed relies on public data retrieved from the CVS repository of the GNOME project and relevant discussion groups. This methodology is described, and results concerning the special characteristics of open source software development are given. These data are used for a first approach to estimating the total effort to be expended.
Structure of Hermes integrations in the germline of the yellow fever mosquito, Aedes aegypti
Authors: Jasinskiene N. , Coates C. J. , James A. A.
Published: March 2003

The Hermes transposable element is derived from the house fly, Musca domestica, and can incorporate into the germline of the yellow fever mosquito, Aedes aegypti. Preliminary Southern analyses indicated that Hermes integrated along with the marker gene into the mosquito genomic DNA. Here we show that Hermes integrations are accompanied by the integration of the donor plasmid as well. In addition, breaks in the donor plasmid DNAs do not occur precisely, or at the end of the terminal inverted repeats, and are accompanied by small deletions in the plasmids. Furthermore, integrations do not cause the typical 8‐bp duplications of the target site DNA. No integrations are observed in the absence of a source of Hermes transposase. The Hermes transposase clearly did not catalyse precise cut‐and‐paste transposition in these transformed lines. It may have integrated the transposon through general recombination or through a partial replicative transposition mechanism. The imprecision of Hermes integration may result from interactions of the transposase with an endogenous hAT‐like element in the mosquito genome.
The mitochondrial genome of the Mediterranean fruit fly, Ceratitis capitata
Authors: Spanos L. , Koutroumbas G. , Kotsyfakis M. , Louis C.
Published: March 2003

The complete sequence of the mitochondrial genome of Ceratitis capitata has been determined. The circular genome is 15 980 bp long and contains a standard gene complement, i.e. the large and small ribosomal RNA subunits, twenty‐two transfer RNA (tRNA) genes and thirteen genes encoding mitochondrial proteins. When comparing the sequence to fragments previously sequenced from other isolates it becomes apparent that interstrain polymorphisms are not rare. These differences are potentially useful for the development of diagnostic tools for population analysis applications, such as determining the source of recent introductions. Moreover, they could help obtain a solution to the long‐lasting controversy on the possible eradication of the Medfly from certain locations.
Two opsin genes from the vetch aphid, Megoura viciae
Authors: Gao N. , Foster R. G. , Hardie J.
Published: March 2003

The cDNAs of two opsins (Megopsin1 and Megopsin2) from the vetch aphid, Megoura viciae, have been sequenced and encoded for gene products with 378 and 371 amino acid residues, respectively. Phylogenetic analysis reveals that Megopsin1 falls into the insect long‐wavelength opsin group and Megopsin2 is a member of the insect UV‐wavelength opsins. Both opsins share the key features of G‐protein‐coupled receptors and the specific motifs of photopigments. In situ hybridization demonstrated that the transcripts of Megopsin1 and Megopsin2 were expressed in the retinula cells of the compound eyes.
Identification of a point mutation in the voltage‐gated sodium channel gene of Kenyan Anopheles gambiae associated with resistance to DDT and pyrethroids
Authors: Ranson H. , Jensen B. , Vulule J. M. , Wang X. , Hemingway J. , Collins F. H.
Published: March 2003

A field trial of permethrin‐impregnated bednets and curtains was initiated in Western Kenya in 1990, and a strain of Anopheles gambiae showing reduced susceptibility to permethrin was colonized from this site in 1992. A leucine–phenylalanine substitution at position 1014 of the voltage‐gated sodium channel is associated with resistance to permethrin and DDT in many insect species, including Anopheles gambiae from West Africa. We cloned and sequenced a partial sodium channel cDNA from the Kenyan permethrin‐resistant strain and we identified an alternative substitution (leucine to serine) at the same position, which is linked to the inheritance of permethrin resistance in the F2 progeny of genetic crosses between susceptible and resistant individuals. The diagnostic polymerase chain reaction (PCR) developed by
Martinez‐Torres et al. [(1998) Insect Mol Biol 7: 179–184] to detect kdr alleles in field populations of An. gambiae will not detect the Kenyan allele and hence reliance on this assay may lead to an underestimate of the prevalence of pyrethroid resistance in this species. We adapted the diagnostic PCR to detect the leucine–serine mutation and with this diagnostic we were able to demonstrate that this kdr allele was present in individuals collected from the Kenyan trial site in 1986, prior to the introduction of pyrethroid‐impregnated bednets. The An. gambiae sodium channel was physically mapped to chromosome 2L, division 20C. This position corresponds to the location of a major quantitative trait locus determining resistance to permethrin in the Kenyan strain of An. gambiae.
Identification of the sibling species of the Anopheles maculipennis complex by heteroduplex analysis
Authors: Romi R. , Boccolini D. , Di Luca M. , La Rosa G. , Marinucci M.
Published: March 2003

The group of anopheline mosquitoes referred to as ‘Anopheles maculipennis complex’ includes the most important malaria vectors of the Palearctic Western region. The species belonging to this complex, however, are difficult or impossible to distinguish by morphological characters. To differentiate sibling palearctic species belonging to this complex, interspecific differences in the ITS2 sequences were used to set up a rapid and sensitive diagnostic tool based on heteroduplex analysis. The relative heteroduplex mobility allowed the following seven species to be readily distinguished: An. atroparvus, An. labranchiae, An. maculipennis s.s., An. martinius, An melanoon, An. messeae and An. sacharovi.
Differential regulation of ribosomal protein gene expression in Aedesaegypti mosquitoes before and after the blood meal
Authors: Niu L. L. , Fallon A. M.
Published: March 2003

In fat body of the mosquito, Aedesaegypti, a cycle of ribosome accumulation and degradation accompanies synthesis of the yolk protein precursor, vitellogenin. Here we compare the transcription and translation of ribosomal proteins rpS6, rpL8 and rpL34, relative to rRNA and vitellogenin genes in Aedesaegypti fat body after eclosion, and in response to a blood meal. Analysis using Northern blots and reverse‐transcription polymerase chain reactions (RT‐PCR) showed that the rpS6, rpL8 and rpL34 genes are coordinately regulated with respect to one another, and that ribosomal protein gene expression in this system was predominantly regulated by transcription during the 3–4 days between adult eclosion and blood feeding. After a blood meal, ribosomal protein mRNA levels remained similar to those in unfed females during the first 18 h, then declined to minimum levels by 48 h after the blood meal. These data indicate that transcription of ribosomal protein genes is low in vitellogenic mosquitoes, relative to previtellogenic females. Experiments with the dissected fat body, however, showed that levels of acetic acid‐soluble proteins increased by approximately threefold between 12 and 24 h after the blood meal. Taken together, these observations suggest that the active translation of ribosomal proteins from stable mRNA accompanies ribosome biosynthesis after the blood meal. Thus, in the fat body of adult female mosquitoes, the expression of ribosomal protein genes is regulated at the level of transcription before the blood meal, while translational control is the predominant regulatory mechanism after the blood meal.
Sequence and organization of the mitochondrial genome of the Chagas disease vector, Triatoma dimidiata
Authors: Dotson E. M. , Beard C. B.
Published: March 2003

The 17 019 bp mitochondrial genome of Triatoma dimidiata is composed of thirteen protein coding sequences, twenty‐two tRNAs, small and large ribosomal units, and a control region. The gene order and orientation are identical to that of Drosophila yakuba. The nucleotide composition is biased toward adenine and thymine (69.5% A + T). The 2.1 kb putative control region, known as the A + T rich region in most insects, has an A + T bias of 66%, but contains a 400 bp sequence that is 77.5% A + T and two other distinct regions: (1) one with a lower A + T bias (60.1%) and (2) a region of eight tandem repeat units.

The identified 1.4 kb nuclear copy of mitochondrial sequences encompasses the string of Gs and the beginning of the cytochrome c oxidase 1 gene but lacks the 1.8 kb region spanning the eight tandem repeats and the 5′ end of the NADH dehydrogenase subunit II gene.
Sindbis virus‐induced silencing of dengue viruses in mosquitoes
Authors: Adelman Z. N. , Blair C. D. , Carlson J. O. , Beaty B. J. , Olson K. E.
Published: March 2003

Aedes aegypti were injected intrathoracically with double subgenomic Sindbis (dsSIN) viruses with inserted sequences derived from the genome of one or more of the four dengue (DEN) virus serotypes. Mosquitoes were highly resistant to challenge with homologous DEN viruses from which the effector sequences were derived, and resistance to DEN viruses was independent of the orientation of the effector RNA. dsSIN viruses designed to express RNA derived from the premembrane coding region of DEN‐2 prevented the accumulation of DEN2 RNA, and C6/36 cells were highly resistant to DEN‐2 virus when challenged at 2, 5 or 8 days after the initial dsSIN virus infections, even though the dsSIN‐derived RNA had sharply declined at the later time points. Initiation of resistance occurred prior to or within the first 8 h after challenge with DEN‐2 virus. We conclude that DEN viruses are inhibited by a mechanism similar to post‐transcriptional gene silencing (PTGS) or RNA interference (RNAi) phenomena described in plants and invertebrates, respectively. The potential occurrence of PTGS or RNAi in mosquitoes and mosquito cells suggests new ways of inhibiting the replication of arthropod‐borne viruses in mosquito vectors, studying vector–virus interactions, and silencing endogenous mosquito genes.
Plant allelochemicals differentially regulate Helicoverpa zea cytochrome P450 genes
Authors: Li Xianchun , Berenbaum May R. , Schuler Mary A.
Published: March 2003

Four cytochrome P450 genes, CYP6B8, CYP6B9, CYP6B27 and CYP6B28, exist in the Helicoverpa zea genome as two pairs of paralogs that evolved from gene duplication and 5′‐polar gene conversion events. RT‐PCR gel blot analyses have shown that all of these genes are expressed constitutively in midguts of all larval instars, suggesting that they have primary roles in the detoxification of plant allelochemicals. Among these, CYP6B9 is expressed only in midgut tissue whereas its paralog, CYP6B27, is expressed primarily in midgut and secondarily in fat body and ovary. CYP6B28 is expressed in midgut, fat body and, to lesser extents in ovary and integument whereas its paralog, CYP6B8, is expressed in midgut and to some extent in fat body. Comparison of the expression levels induced by eight plant allelochemicals, one drug (phenobarbital), and an insecticide (α‐cypermethrin) indicates that, for the most part, the four P450s respond individually to these inducers, with all four induced strongly by chlorogenic acid, a shikimate pathway intermediate and a lignin biosynthesis intermediate present in a wide variety of plants, and indole‐3‐carbinol, a glucobrassicin breakdown product present in the Brassicaceae. The multiple levels at which these P450 genes are apparently diverging (e.g. transcriptional responses, protein sequences) support the suggestion that gene conversion events facilitate gene evolution by allowing duplicated copies greater time to acquire selectable differences in both coding and promoter sequences.
Comparison of EST libraries from seven beetle species: towards a framework for phylogenomics of the Coleoptera
Authors: Theodorides K. , De Riva A. , Gómez‐Zurita J. , Foster P. G. , Vogler A. P.
Published: March 2003

Relatively little is known about Coleoptera genes and genomes and how these compare in different taxa. We describe here the construction, DNA sequencing and sequence comparisons of cDNA libraries from seven beetle species. A total of 6717 bacterial colonies were screened for cDNA insert containing plasmids and 2784 size selected clones were 5′‐ and 3′‐end sequenced to produce 1620 assembled sequences. Similarity comparisons with existing protein sequence databases revealed that 65.1% had matches (E < 10−4) in other organisms, with greater numbers of matches in Drosophila melanogaster than Caenorhabditis elegans and Saccharomyces cerevisiae databases. tBlastX comparisons also revealed numerous similarity hits (E < 10−20) in intra‐ and interlibrary comparisons. These results show the potential of small cDNA libraries for discovery and comparative analysis of genes useful for phylogenomic and functional studies.
Comparative study of retreatment of Thermafil and lateral condensation endodontic fillings
Authors: Frajlich  , Goldberg  , Massone  , Cantarini  , Artaza
Published: March 2003

The aim of the present study was to evaluate the possibility of retreating teeth obturated with: (i) Thermafil with plastic carrier, (ii) Thermafil with metallic carrier and (iii) laterally condensed gutta‐percha. The following aspects were analysed: removal of obturation, time required for removal, remaining filling material and apical extrusion during reinstrumentation. Thirty human teeth with a single canal were instrumented up to a size 45 K‐type file, divided into three groups of 10 teeth each and obturated with Thermafil with plastic carrier (group 1), Thermafil with metallic carrier (group 2), and laterally condensed gutta‐percha (group 3) with AH26 as the sealer. Reinstrumentation was performed manually after 30 days storage using Hedstroem files and xylene as a solvent. The average time needed to remove the obturation was 12 min 1 s for group 1, 14 min 36 s for group 2 and 11 min 26 s for group 3. Although no statistically significant differences were observed between groups 1 and 2, and 1 and 3 (P > 0.05), the differences between groups 2 and 3 were significant (P < 0.05). The amount of remaining filling material was significantly higher in group 2 when compared with groups 1 and 2 (P < 0.01). Although the lowest incidence of apical extrusion during reinstrumentation was found in group 3, the differences among groups were not statistically significant.
Endodontic retreatment decisions: no consensus
Authors: Aryanpour S. , Van Nieuwenhuysen J. ‐P. , D'Hoore W.
Published: March 2003

Aim  The objectives of the present study were to: (i) evaluate the consensus, if any, amongst dental schools, students and their instructors managing the same clinical cases, all of which involved endodontically treated teeth; and (ii) determine the predominant proposed treatment option.

Methodology  Final year students, endodontic staff members and instructors of 10 European dental schools were surveyed as decision makers. Fourteen different radiographic cases of root canal treated teeth accompanied by a short clinical history were presented to them in a uniform format. For each case the decision makers were requested to: (i) choose only one out of nine treatment alternatives proposed, from ‘no treatment’ to ‘extraction’ via ‘retreatment’ and ‘surgery’ (ii) assess on two 5‐point scales: the difficulty of making a decision, and the technical complexity of the retreatment procedure.

Results  The results indicate wide inter‐ and also intra‐school disagreements in the clinical management of root canal treated teeth. Analysis of variance showed that the main source of variation was the ‘school effect’, explaining 1.8% (NS) to 18.6% (P< 0.0001) of the treatment variations. No other factor explained as much variance. Decision difficulty was moderately correlated to technical complexity (Pearsons' r ranging from 0.19 to 0.35, P< 0.0001).

Conclusions  No clear consensus occurred amongst and within dental schools concerning the clinical management of the 14 cases. The lack of consensus amongst schools seems to be due mainly to chance or uncertainty, but can be partly explained by the ‘school effect’.
Influence of calcium hydroxide intracanal dressings on the prognosis of teeth with endodontically induced periapical lesions
Authors: Weiger R. , Rosendahl R. , Löst C.
Published: March 2003

Aim This prospective clinical study explored the influence of calcium hydroxide as an interappointment dressing on the healing of periapical lesions associated with pulpless teeth that had not been endodontically treated previously. This was achieved by comparing the prognosis after a two‐visit root canal treatment with that following a one‐visit treatment.

Methodology Seventy‐three patients were recruited having one tooth with an endodontically induced lesion. Of these patients, 67 could be re‐examined. Calcium hydroxide was placed in the instrumented root canals of 31 teeth for at least one week and the treatment finished at the second visit. Thirty‐six teeth were root canal treated at one visit. The criteria for success were the absence of signs and symptoms indicating an acute phase of periapical periodontitis and radiographically a periodontal ligament space ofnormal width.

Methods for event time analysis were used to evaluate and compare the prognosis of both treatment approaches.

Results The probability that complete periapical healing will take place increased continuously with the length of the observation period. In both treatment groups the likelihood that the root canal treatment yields a success within an observation time of five years exceeded 90%. A statistically significant difference between the two treatment groups could not be detected.

Conclusions From a microbiological perspective, one‐visit root canal treatment created favourable environmental conditions for periapical repair similar to the two‐visit therapy when calcium hydroxide was used as antimicrobial dressing. One‐visit root canal treatment is an acceptable alternative to two‐visit treatment for pulpless teeth associated with an endodontically induced lesion.
Root canal treatment in general practice in Sudan
Authors: Ahmed M. F. , Elseed A. I. , Ibrahim Y. E.
Published: March 2003

Aim

The aim of this study was to evaluate the practice and depth of knowledge of root canal treatment by dental practitioners in Khartoum, the capital city of Sudan, in order to improve the current status of endodontic therapy.

Methodology

A questionnaire was posted to 55 registered dental practitioners. Completed questionnaires were analysed in term of simple summary statistics.

Results

A total of fifty‐two (95%) practitioners responded. Eighty‐five per cent of the respondents indicated that they performed root canal treatment for their patients. Of these, 84% included molars in their activity. Amongst those who carried out root canal treatment, only one practitioner used rubber dam for isolation, whilst the remainder used cotton wool rolls. The majority of respondents (80%) used hydrogen peroxide to irrigate canals during treatment. Three‐quarters of practitioners used formocresol as an interappointment medicament. The stepback preparation technique was the method of choice for 98% of respondents. All practitioners used hand instruments to prepare root canals and all used gutta‐percha for obturation; three‐quarters of them used cold lateral condensation for all or some cases. The average number of radiographs routinely taken for root canal treatment was three. Only 73% used radiographs for measuring the working length. Ninety‐five per cent of respondents indicated that they usually completed a root filling in three or more visits. Three‐quarters of practitioners restored the teeth permanently immediately after the obturation and one‐quarter preferred waiting for 1 or 2 weeks.

Conclusions

In Sudan, there are no dental practices limited to endodontics and no postgraduate training programmes. This survey shows the importance of establishing higher specialist training or continuing dental education for practitioners to update their knowledge.
Use of rubber dam and irrigant selection in UK general dental practice
Authors: Whitworth J. M. , Seccombe G. V. , Shoker K. , Steele J. G.
Published: March 2003

Aim

To evaluate factors which influence rubber dam use and irrigant selection in UK National Health Service (NHS) endodontics.

Methodology

A postal survey was conducted amongst two age cohorts of dentists, representing all of the 1970–73 (older) and 1990–93 (younger) graduates of two northern English dental schools (n = 643). Key and supplementary questions were posed on levels of rubber dam use, irrigant selection, and factors influencing practice in NHS endodontics. After manual checking, validated (dual) entry of responses was made to a flat ASCII data file before analysis with SPSS software. The threshold for statistical significance was set at the 95% probability level.

Results

Eighty‐five per cent of the valid sample responded to the questionnaire. Regardless of age and qualifying school, less than one‐fifth of dentists always or frequently used rubber dam, whilst 60% never used it. Qualifying school had a significant influence on rubber dam use, whilst age had a variable influence. Major disincentives to the use of rubber dam included the perception that patients do not like it, that the NHS fee was inadequate to justify its use, that it took too long to apply, and that dentists had received inadequate training. Frequent users of rubber dam were significantly less likely to cite these disincentives than nonusers. Overall, local anaesthetic solution was the most common endodontic irrigant. Irrigant choice was strongly linked to rubber dam use, and to graduation cohort. Seventy‐one per cent of rubber dam users irrigated with sodium hypochlorite, compared with only 38% of nonusers. This pattern was reversed for local anaesthetic irrigation. Younger graduates were significantly more likely to irrigate with local anaesthetic solution than their older counterparts, and the younger graduates of one school showed a highly significant increase in the use of chlorhexidine.

Conclusions

1) The majority of UK Health Service dentists never use rubber dam isolation in endodontic treatment. 2) Qualifying school has a significant impact on rubber dam use, and irrigant selection. 3) Use of rubber dam has a significant association with irrigant choice in endodontics.
The efficacy of gutta‐percha removal using ProFiles
Authors: Ferreira J. J. , Rhodes J. S. , Pitt Ford T. R.
Published: March 2003

Aim

The purpose of this study was to compare the efficacy in vitro of gutta‐percha removal from obturated root canals using ProFiles.

Methodology

Forty‐eight human root canals with curvatures ranging between 25 and 45° were instrumented by a standardized method to an apical ISO size 30 and 0.04 taper. They were obturated with vertically condensed gutta‐percha. Retreatment was performed with the following techniques: K‐Flexofiles with chloroform; Hedstrom files with chloroform; ProFiles 0.04 taper with chloroform; ProFiles 0.04 taper alone. The time for each method was measured. A microfocal macroradiographic technique was used to evaluate the amount of debris remaining within the root canals after the retreatment procedure. Roots were divided into apical, middle and coronal parts and scored on a scale of 0 (no debris) to 3 (>50% of walls covered with debris) by trained observers on two separate occasions.

Results

The scores for debris remaining within root canals for K‐Flexofiles with chloroform and ProFiles with chloroform were the lowest and not significantly different at all three levels of the roots examined (P > 0.05), and Hedstrom files with chloroform and ProFiles with chloroform were not significantly different in the apical part. In general, coronal parts were cleaner than apical parts. The difference in scores at the three levels between ProFiles with chloroform and ProFiles alone were each significant (P < 0.01). Instrumentation using ProFiles with chloroform (mean 6.42 min) was significantly faster than using hand files (mean 11.67 min) (P < 0.01).

Conclusion

The results indicated that ProFiles or hand files with chloroform produced similarly clean canals, but that ProFiles were faster.
A methodology for quantitative evaluation of root canal instrumentation using microcomputed tomography
Authors: Bergmans L. , Van Cleynenbreugel J. , Wevers M. , Lambrechts P.
Published: March 2003

Aim

The aim of this paper was to present an objective methodology for quantitative evaluation of root canal instrumentation using microcomputer tomography, together with developed software based on a constructed mathematical model.

Methodology

A desktop X‐ray micro‐CT scanner (SkyScan 1072) was used to provide data sets of an extracted mandibular molar before and after instrumentation (ProFile 0.04 Taper instruments, Dentsply Maillefer, Ballaigues, Switzerland) that were stored for later use by software. A volume visualization package (T3D) was applied to obtain 3D renderings of the molar to illustrate the qualitative visualization capacity. Next, medical image volume fusion software was used to allow alignment of pre and post image volumes. Finally, software was developed to make quantitative measurements and to provide additional qualitative information on the registered image volumes. This procedure implemented a true 3D mathematical model for quantification of instrumentation effects. Using a local co‐ordinate frame, perpendicular reslices were made at five different levels within the mesio‐buccal canal to evaluate transportation and centring ability.

Results

At first accurate and detailed 3D renderings were obtained. The general and local canal shape before and after preparation could visually be examined in 360° rotation. Numerical values were obtained for volume, volume changes and transportation. Centring ability was calculated by centring ratio and centre movement.

Conclusions

This methodology is a new and objective way for quantitative evaluation of root canal instrumentation using microcomputer tomography and dedicated software.
Root canal treatment in an unusual maxillary first molar: a case report
Authors: Fava L. R. G.
Published: March 2003

Aim

The aim of this clinical article is to describe the unusual anatomy that was detected in a maxillary first molar during routine endodontic treatment.

Success in root canal treatment is achieved after thorough cleaning and shaping followed by the complete obturation of the root canal system. Such treatment may be performed in root canal systems that do not comply with the normal anatomical features described in standard textbooks. The present case describes root canal treatment in a maxillary first molar with two roots and a type IV canal configuration in the buccal root.

Key learning points

Careful examination of radiographs and the internal anatomy of teeth is essential.

Root canal treatment is likely to fail if the entire system is not debrided and filled.

Anatomic variations can occur in any tooth.
Relationship between number of proximal contacts and survival of root canal treated teeth
Authors: Caplan D. J. , Kolker J. , Rivera E. M. , Walton R. E.
Published: March 2003

Aim The present study tested the hypothesis that having two proximal contacts (PCs) at access is associated with improved survival of root canal treated (RCT) teeth, controlling for important presenting conditions, endodontic variables and restorative factors.

Methodology A treatment database at the University of Iowa College of Dentistry was used to identify permanent teeth undergoing obturation between 1 July 1985 and 31 December 1987. The list was restricted to teeth of patients with at least one dental visit in each two‐year interval from 1985 to 1996, and a simple random sample of 280 patients (n = 400 teeth) was selected. Dental charts, radiographs, and computerized databases were examined to ascertain variables of interest and to verify study inclusion criteria. Kaplan–Meier survival estimates were generated for the 221 teeth satisfying study inclusion criteria. Multivariate Cox models were developed, with standard errors adjusted to account for clustering of teeth within patients.

Results The final Cox model showed that teeth with ≤1 PC at access were lost at a rate three times that of teeth with 2 PCs (hazard ratio = 3.1; 95% confidence interval = 1.9–5.1), controlling for tooth type, presence of radiographic caries at access, and presence of a crown before or after obturation.

Conclusions Because RCT teeth with two PCs at access experienced substantially better survival than teeth with fewer than two PCs, the influence of PCs on prognosis should be recognized during treatment planning. Future research should employ prospective study designs, capture additional variables, and provide data to support endodontic treatment decisions.
A comparison of the shaping characteristics of two nickel–titanium endodontic hand instruments
Authors: Dobó‐Nagy C. , Serbán T. , Szabó J. , Nagy G. , Madléna M.
Published: March 2003

Aim The purpose of this study was to compare the shaping characteristics of Ni–Ti K‐files and Ni–Ti S‐files manipulated by hand.

Methodology A total of 60 extracted human roots were embedded in resin blocks. The embedded roots were divided into three groups: (i) roots with straight; (ii) apically curved; and (iii) continuously curved canals. Each of the three groups was randomly divided into two subgroups; one subgroup in each group was prepared with Ni–Ti K‐files and the other with Ni–Ti S‐files. The files were used with a step‐back technique and enlarged so that the master apical file was size 30, and the canals were stepped back to size 40. The performance of the files was assessed by the superimposition of projected radiographs taken in bucco‐lingual and mesio‐distal directions before and after the preparation. The results were analysed statistically using analysis of covariance and Duncan’s multiple range test.

Results Although canal preparation using Ni–Ti K‐files was quicker, there were no statistically significant differences between file types. The Ni–Ti S‐file removed significantly more material at the most coronal level (P < 0.05). There were minor differences between instruments at the apical level. Only in the proximal view of apically curved canals prepared with Ni–Ti S‐files was significantly more dentine removed from the outer aspect of the curvature (P < 0.05). At the middle level (wide danger zone) the Ni–Ti S‐files removed more dentine from the inner aspect of the curvature in those roots with apically curved canals.

Conclusions Under the conditions of this study, preparation with Ni–Ti K‐files produced more appropriate shapes in roots with apically curved canals than Ni–Ti S‐files.
Mechanisms involved in the resistance of Enterococcus faecalis to calcium hydroxide
Authors: Evans M. , Davies J. K. , Sundqvist G. , Figdor D.
Published: March 2003

Aim This study sought to clarify the mechanisms that enable E. faecalis to survive the high pH of calcium hydroxide.

Methodology E. faecalis strain JH2‐2 was exposed to sublethal concentrations of calcium hydroxide, with and without various pretreatments. Blocking agents were added to determine the role of stress‐induced protein synthesis and the cell wall‐associated proton pump.

Results E. faecalis was resistant to calcium hydroxide at a pH of 11.1, but not pH 11.5. Pre‐treatment with calcium hydroxide pH 10.3 induced no tolerance to further exposure at pH 11.5. No difference in cell survival was observed when protein synthesis was blocked during stress induction, however, addition of a proton pump inhibitor resulted in a dramatic reduction of cell viability of E. faecalis in calcium hydroxide.

Conclusions Survival of E. faecalis in calcium hydroxide appears to be unrelated to stress induced protein synthesis, but a functioning proton pump is critical for survival of E. faecalis at high pH.
To store or mature spermatozoa? The primary role of the epididymis
Authors: Jones
Published: March 2003

The function of the epididymis is considered in the context that it is necessary to transport spermatozoa for internal fertilization, but is also an adaptation involved in the competition between males to achieve paternity. Post‐testicular sperm maturation and storage occur in lower vertebrates, such as the chondrichthyes, in which sperm storage is essential due to the slow rate of spermatogenesis. These roles persist in higher vertebrates, including mammals. However, they are less important in certain birds, in which the rate of sperm production by the testes is sufficient to supply enough spermatozoa to cope with the demands of a competitive mating system.

There is no evidence for mammals, other than humans, that spermatozoa can achieve the capacity to fertilize an ovum without passing through some of the proximal epididymis. Storage of spermatozoa in the epididymis is probably not essential for a male to achieve conception in a protected mating system, but is very important in a competitive mating system. There is some variation between species in the magnitude of the epididymal storage region. This is related to the descent of the epididymides (and testes) into a scrotal sac in some species and/or the demands of the mating system in other species. The claims that humans are not dependent on post‐testicular sperm maturation and storage need to be qualified, as they are based on studies of abnormal tracts and tests of fertility which are not rigorous. Nevertheless, the claims are of considerable clinical significance and may indicate that humans are less dependent on post‐testicular sperm maturation and storage than other mammals. This may be an adaptation of the testes and epididymides to the scrotal conditions of modern man or a response to changing environmental factors.
Co‐localization of HP1 and TP1 transcripts in human spermatids by double electron microscopy in situ hybridization
Authors: Siffroi  , Alfonsi  , Dadoune
Published: March 2003

Nuclear changes in the basic nucleoprotein complement occur during spermiogenesis in man. Somatic type histones are displaced by transition proteins which are replaced themselves by protamines, the major nuclear proteins present in late spermatids and sperm nuclei. Sense and antisense 35S‐labelled riboprobes, coding respectively for human transition protein 1 (TP1) and protamine 1 (HP1), were synthesized with modified specific oligonucleotides and were used for light microscopy in situ hybridization. A double EM in situ hybridization was performed using a digoxigenin‐labelled probe for TP1 and a biotin‐labelled probe for HP1, and hybrids were revealed, respectively, with specific antibodies coupled to colloidal gold particles of different sizes (10 nm and 15 nm). For both types of transcripts, histological study revealed a specific distribution of the silver grains in the adluminal region of the seminiferous tubules where spermatids are localized. Quantitative ultrastructural analysis of the nuclear and cytoplasmic labelling densities for the mRNAs coding for TP1 and HP1 showed that the transcripts were found in both the nucleus and cytoplasm of round spermatids and persisted until the elongation phase. Transcripts accumulated in the spermatid cytoplasm without any particular cellular compartmentalization. At the end of the spermatid elongation phase, the disappearance of TP1 and HP1 transcripts may be related to the arrest of transcriptional activity, while the deposition of transition proteins and protamines occurs successively within spermatid nuclei.
Relative contribution of leukocytes and of spermatozoa to reactive oxygen species production in human sperm suspensions
Authors: Whittington Kate , Ford
Published: March 2003

The contribution of leukocytes and of spermatozoa to reactive oxygen species (ROS) production in prepared sperm suspensions from donors and subfertility patients was compared. In both groups, more leukocytes/106 spermatozoa were counted in samples which produced detectable ROS than in those that did not: Donors – 645 vs. 170 (medians, n = 7; p < 0.01, Kruskal–Wallis), Subfertile group – 1785 (n = 18) vs. 11 (n = 8) (p < 0.005, Kruskal–Wallis), respectively. Leukocyte concentrations were correlated with basal (r = 0.826, p < 0.001) and with ROS production stimulated with 50 μmol N‐formyl, met, leu, phe l−1 (N‐FMLP) (r = 0.835, p < 0.001) and 100 nmol phorbol 12‐myristate 13‐acetate l−1 (PMA) (r = 0.835, p < 0.001) measured using a chemiluminescence assay. Leukocytes were removed from the sperm suspensions of 6 donors and from 96 ejaculates from 21 subfertility patients and ROS production was determined. Subsequently, in all 6 donors, N‐FMLP did not stimulate ROS production indicating that leukocyte removal was complete, though in one case PMA stimulated low levels of ROS production. In 65 ejaculates from subfertile men the N‐FMLP response was completely eliminated but in 7 of these samples PMA continued to stimulate ROS production. We conclude that infiltrating leukocytes are the predominant source of ROS production in unpurified sperm preparations. Some purified sperm suspensions could be stimulated to produce ROS by the addition of PMA indicating that spermatozoa themselves may produce ROS, albeit in much smaller amounts.
Biphasic effect of nitric oxide on testosterone and cyclic GMP production by purified rat Leydig cells cultured in vitro
Authors: Valenti Sandra , Cuttica Carla Micaela , Fazzuoli Laura , Giordano Giulio , Giusti Massimo
Published: March 2003

Nitric oxide (NO) biphasically modulates osteoclast function and sperm motility by exerting a positive effect at low concentrations and a negative effect at high concentrations. We therefore tested whether NO exerts a comparable effect on testosterone secretion by cultured rat Leydig cells. Three NO‐donors, S‐nitroso‐ N‐acetylpenicillamine (SNAP), diethylamine/nitric oxide complex sodium salt (DEA/NO) and diethylenetriamine nitric oxide adduct (DETA/NO) were administered in a wide range of concentrations (10–8–10–3 M for 3 h) to Percoll‐purified Leydig cells from adult rats. These drugs raised testosterone and cGMP secretion when used at low concentrations (10–8–10–5 M); however, they inhibited testosterone, but did not affect cGMP, secretion at concentrations higher than 10–5 M. Administration of the NO scavenger haemoglobin (160 μg/mL) prevented both the stimulatory and the inhibitory effect of these drugs. Nitrite accumulation was measured as a marker of NO released by the drugs in our in vitro system; it fell within the range of control media in the presence of NO‐donor concentrations lower than 10–5 M, but was several‐fold higher in the media of cells treated with concentrations of the NO‐donors greater than 10–5 M. These data show that (1) NO exerts a biphasic effect on testosterone secretion, which is stimulatory at low and inhibitory at high concentrations; (2) the stimulatory effect of NO is mediated by cGMP, the classic second messenger for NO action.
Chlorinated pesticides and heavy metals in human semen
Authors: Kumar  , Pant  , Srivastava
Published: March 2003

The concentration of chlorinated pesticides and heavy metals (lead and cadmium) was measured using gas liquid chromatography and the graphite tube atomizer of atomic absorption spectrophotometer, respectively, in semen samples collected from men in the normal human population. Significant concentrations of lead and cadmium were detected. Significant amounts of hexachlorocyclohexane (HCH) and its isomers alpha, beta, gamma and delta, the dichlorodiphenyl trichloroethane (DDT) metabolite 1,1,1‐trichloro‐2,2‐bis (p chlorophenyl ethane) (pp′‐DDE) and low values of 1,1,2‐dichloro‐2,2‐bis(p chlorophenyl ethane) (pp′‐DDD) aldrin or endosulfan were detected. The presence of these xenobiotics in human semen might be related to the extensive use of pesticides, emission of exhaust from motor vehicles, consumption of tobacco and industrial operations.
Semen quality over a 10‐year period in 22,249 men in Korea
Authors: Seo Ju Tae , Rha Koon Ho , Park Yong‐Seog , Lee Moo Sang
Published: March 2003

A retrospective study was conducted in a large population presenting with infertility to determine whether sperm quality has changed in Korea in the last 10 years. We reviewed sperm concentration, motility and semen volume in 22,249 men from whom semen was collected in our laboratory between January 1989 and April 1998 and analysed according to WHO (1987) guidelines. Mean age of the men was 32 years (range 21–40). Data were collected in healthy men with infertility. The mean sperm concentration was 60.5 ×106/mL from 1989 to 1998. There was no statistically significant difference for each year (p > 0.05). Semen volume and sperm motility were also unchanged during the same time period. There was no significant association between either age or year of birth and semen quality. Of the total population, 4033 men (19.0%) exhibited azoospermia and 8397 men (40.1%) had normal semen parameters which satisfied the 1987 WHO criteria. The changes observed in the semen parameters analysed in this large population showed no evidence of deteriorating semen quality in Korea over the last 10 years.
Role of genitourinary inflammation in infertility: synergistic effect of lipopolysaccharide and interferon‐γ on human spermatozoa
Authors: Sikka Suresh C. , Champion Hunter C. , Bivalacqua Trinity J. , Estrada Lance S. , Wang Run , Rajasekaran Mahadevan , Aggarwal Bharat B. , Hellstrom Wayne J. G.
Published: March 2003

Pro‐inflammatory cytokines are elevated in the semen of patients with genitourinary inflammation (GUI). Whether this increase in cytokines in GUI patients plays any critical role in male factor infertility is not clear. The present study investigated the in vitro effects of two important pro‐inflammatory cytokines, lipopolysaccharide (LPS) and interferon‐γ (IFN‐γ), on sperm motility, viability, membrane integrity and motion parameters. Washed spermatozoa from healthy donors were incubated with LPS (0.1 mg/mL) or IFN‐γ (0.1 mg/mL) alone or in combination. Sperm motility, viability, membrane integrity and computer‐assisted motion were evaluated at various time intervals (0, 30, 60 and 180 min) after treatment. Sperm membrane integrity was analysed using the hypo‐osmotic swelling test (HOST). LPS and IFN‐γ individually did not alter sperm viability or motility, but their combination showed a significant time‐dependent decrease (p < 0.05) in sperm motility, viability and membrane integrity. Sperm motion parameters (straight‐line velocity, curvilinear velocity, mean linearity, or amplitude of lateral head displacement) were not affected by LPS or IFN‐γ at the concentrations used in this study. These data suggest that the combination of LPS and IFN‐γ is detrimental to human spermatozoa and may contribute to male factor infertility in patients with chronic GUI.
Effect of pentoxifylline on motility and membrane integrity of cryopreserved human spermatozoa
Authors: Stanic P. , Sonicki Z. , Suchanek E.
Published: March 2003

The purpose of this study was to examine the effects of pentoxifylline used before and after semen cryopreservation–thawing on sperm motility and membrane integrity. Twenty‐four semen samples were split into four equal aliquots. Aliquots were incubated at 37 °C for 30 min, followed by cryopreservation with TEST‐yolk freezing medium using slow programmable freezing protocol. After 2 weeks the sperm samples were thawed, washed twice in Quinn's Sperm Washing Medium (modified HTF with 5.0 mg/mL Human Albumin) and incubated at 37 °C for 30 min. Aliquots were treated by adding 3 mmol/L pentoxifylline to: (1) fresh sperm samples during incubation period prior to cryopreservation, (2) sperm samples as a supplement to the cryoprotectant prior to cryopreservation, and (3) thawed sperm samples during incubation period. One aliquot received no treatment (control group). The addition of 3 mmol/L pentoxifylline to fresh semen during incubation period prior to cryopreservation significantly decreased progressive and total motility compared with controls. However, the addition of 3 mmol/L pentoxifylline to cryopreserved semen after thawing significantly increased progressive and total motility compared with controls. After post‐thaw, no differences in motion characteristics between sperm samples treated by adding 3 mmol/L pentoxifylline as a supplement to the cryoprotectant and control groups were observed. Post‐thaw hypoosmotic swelling (HOS) test scores did not improve with the addition of pentoxifylline compared with the control group. It is concluded that pentoxifylline enhanced post‐thaw motility of cryopreserved human spermatozoa when added after thawing. No improvement was found by freezing sperm with pentoxifylline.
Role of seminal osmolarity in the regulation of human sperm motility
Authors: Rossato M. , Balercia G. , Lucarelli G. , Foresta C. , Mantero F.
Published: March 2003

Mammalian sperm at ejaculation are suspended in the seminal plasma, a heterogeneous mixture deriving from the testicular/epididymal fluid and from secretions of seminal vesicles, prostate and bulbourethral glands. Biochemical characteristics of seminal fluid change along the male reproductive tract when considering its inorganic and organic composition and pH but it is known that in each region of the male genital tract seminal osmolarity is higher than that of serum. It has been previously demonstrated that in invertebrate and vertebrate sperm, seminal plasma osmolarity influences sperm motility and activity, and human sperm have been shown to possess osmosensitive calcium entry pathway that controls important functions such as acrosome reaction and oocyte penetration. In the present study, we have determined seminal plasma osmolarity in a large number of normozoospermic fertile and asthenozoospermic infertile subjects correlating it with sperm motility percentages and kinematic characteristics determined utilizing a computerized motion analysis system. Our results confirm that seminal plasma osmolarity is higher than that of serum (336.1 ± 20.2 vs. 291.1 ± 6.9 mOsm/L, respectively). Normozoospermic subjects show seminal osmolarity values that are significantly lower with respect to asthenozoospermic patients (317.8 ± 12.2 vs. 345.2 ± 22.6 mOsm/L, p < 0.001), irrespective of the cause of asthenozoospermia. Seminal plasma osmolarity negatively correlates with sperm progressive motility percentages and kinetic characteristics (curvilinear velocity, linear velocity, linear coefficient and lateral displacements of sperm head). Furthermore, when sperm from fertile subjects were suspended in medium with an osmolarity increasing from 300 to 600 mOsm, sperm motility percentages and kinetics characteristics were progressively reduced and nearly abolished when medium osmolarity was 600 mOsm. On the contrary, when sperm from asthenozoospermic subjects with high semen osmolarity were resuspended in medium with lower osmolarity, sperm motility parameters improved significantly. Sperm motility parameters did not correlate with seminal plasma concentrations of sodium, potassium, chloride with a weak correlation only with seminal calcium concentration. No correlations are present between seminal plasma osmolarity and ionic composition.

In conclusion, the present study confirms and extends the knowledge that, in human, seminal plasma osmolarity is higher than that of serum and demonstrates that seminal osmolarity influences sperm motility characteristics and then it may contribute to the pathogenesis of some forms of asthenozoospermia and male infertility.