RNA profiling of human testicular cells identifies syntenic lncRNAs associated with spermatogenesis.

STUDY QUESTION: Is the noncoding transcriptional landscape during spermatogenesis conserved between human and rodents? SUMMARY ANSWER: We identified a core group of 113 long noncoding RNAs (lncRNAs) and 20 novel genes dynamically and syntenically transcribed during spermatogenesis. WHAT IS KNOWN ALREADY: Spermatogenesis is a complex differentiation process driven by a tightly regulated and highly specific gene expression program. Recently, several studies in various species have established that a large proportion of known lncRNAs are preferentially expressed during meiosis and spermiogenesis in a testis-specific manner. STUDY DESIGN, SIZE, DURATION: To further investigate lncRNA expression in human spermatogenesis, we carried out a cross-species RNA profiling study using isolated testicular cells. PARTICIPANTS/MATERIALS, SETTING, METHODS: Human testes were obtained from post-mortem donors (N = 8, 51 years old on average) or from prostate cancer patients with no hormonal treatment (N = 9, 80 years old on average) and only patients with full spermatogenesis were used to prepare enriched populations of spermatocytes, spermatids, Leydig cells, peritubular cells and Sertoli cells. To minimize potential biases linked to inter-patient variations, RNAs from two or three donors were pooled prior to RNA-sequencing (paired-end, strand-specific). Resulting reads were mapped to the human genome, allowing for assembly and quantification of corresponding transcripts. MAIN RESULTS AND THE ROLE OF CHANCE: Our RNA-sequencing analysis of pools of isolated human testicular cells enabled us to reconstruct over 25 000 transcripts. Among them we identified thousands of lncRNAs, as well as many previously unidentified genes (novel unannotated transcripts) that share many properties of lncRNAs. Of note is that although noncoding genes showed much lower synteny than protein-coding ones, a significant fraction of syntenic lncRNAs displayed conserved expression during spermatogenesis. LARGE SCALE DATA: Raw data files (fastq) and a searchable table (.xlss) containing information on genomic features and expression data for all refined transcripts have been submitted to the NCBI Gene Expression Omnibus under accession number GSE74896. LIMITATIONS, REASONS FOR CAUTION: Isolation procedures may alter the physiological state of testicular cells, especially for somatic cells, leading to substantial changes at the transcriptome level. We therefore cross-validated our findings with three previously published transcriptomic analyses of human spermatogenesis. Despite the use of stringent filtration criteria, i.e. expression cut-off of at least three fragments per kilobase of exon model per million reads mapped, fold-change of at least three and false discovery rate adjusted P-values of less than <1%, the possibility of assembly artifacts and false-positive transcripts cannot be fully ruled out. WIDER IMPLICATIONS OF THE FINDINGS: For the first time, this study has led to the identification of a large number of conserved germline-associated lncRNAs that are potentially important for spermatogenesis and sexual reproduction. In addition to further substantiating the basis of the human testicular physiology, our study provides new candidate genes for male infertility of genetic origin. This is likely to be relevant for identifying interesting diagnostic and prognostic biomarkers and also potential novel therapeutic targets for male contraception. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by l'Institut national de la santé et de la recherche médicale (Inserm); l'Université de Rennes 1; l'Ecole des hautes études en santé publique (EHESP); INERIS-STORM to B.J. [N 10028NN]; Rennes Métropole 'Défis scientifiques émergents' to F.C (2011) and A.D.R (2013). The authors have no competing financial interests.

Extracorporeal circuit for Panton-Valentine leukocidin-producing Staphylococcus aureus necrotizing pneumonia.

OBJECTIVE: To describe two cases of Panton-Valentine leukocidin-producing Staphylococcus aureus (PVL-SA) necrotizing pneumonia treated with ECMO, and complete pulmonary evaluation at six months. METHODS: Retrospective analysis of two patients presenting with severe PVL-SA pneumonia who both underwent complete respiratory function testing and chest CT scan six months after hospital discharge. RESULTS: Indications for ECMO were refractory hypoxia and left ventricular dysfunction associated with right ventricular dilatation. Patients were weaned off ECMO after 52 and 5 days. No ECMO-related hemorrhagic complication was observed. Pulmonary function tests performed at six months were normal and the CT scan showed complete regression of pulmonary injuries. CONCLUSION: PVL-SA pneumonia is characterized by extensive parenchymal injuries, including necrotic and hemorrhagic complications. ECMO may be used as a salvage treatment without any associated hemorrhagic complication, provided anticoagulant therapy is carefully monitored, and may lead to complete pulmonary recovery at six months.

Characteristics and prognosis of pneumococcal endocarditis: a case-control study.

Case series have suggested that pneumococcal endocarditis is a rare disease, mostly reported in patients with co-morbidities but no underlying valve disease, with a rapid progression to heart failure, and high mortality. We performed a case-control study of 28 patients with pneumococcal endocarditis (cases), and 56 patients with non-pneumococcal endocarditis (controls), not matched for sex and age, during the years 1991-2013, in one referral centre. Alcoholism (39.3% versus 10.7%; p <0.01), smoking (60.7% versus 21.4%; p <0.01), the absence of previously known valve disease (82.1% versus 60.7%; p 0.047), heart failure (64.3% versus 23.2%; p <0.01) and shock (53.6% versus 23.2%; p <0.01) were more common in pneumococcal than in non-pneumococcal endocarditis. Cardiac surgery was required in 64.3% of patients with pneumococcal endocarditis, much earlier than in patients with non-pneumococcal endocarditis (mean time from symptom onset, 14.1 ± 18.2 versus 69.0 ± 61.1 days). In-hospital mortality rates were similar (7.1% versus 12.5%). Streptococcus pneumoniae causes rapidly progressive endocarditis requiring life-saving early cardiac surgery in most cases.

Severe epilepsy as the major symptom of new mutations in the mitochondrial tRNA(Phe) gene.

OBJECTIVE: To present 2 families with maternally inherited severe epilepsy as the main symptom of mitochondrial disease due to point mutations at position 616 in the mitochondrial tRNA(Phe) (MT-TF) gene. METHODS: Histologic stainings were performed on skeletal muscle slices from the 2 index patients. Oxidative phosphorylation activity was measured by oxygraphic and spectrophotometric methods. The patients' complete mitochondrial DNA (mtDNA) and the relevant mtDNA region in maternal relatives were sequenced. RESULTS: Muscle histology showed only decreased overall COX staining, while a combined respiratory chain defect, most severely affecting complex IV, was noted in both patients' skeletal muscle. Sequencing of the mtDNA revealed in both patients a mutation at position 616 in the MT-TF gene (T>C or T>G). These mutations disrupt a base pair in the anticodon stem at a highly conserved position. They were apparently homoplasmic in both patients, and had different heteroplasmy levels in the investigated maternal relatives. CONCLUSIONS: Deleterious mutations in the mitochondrial tRNA(Phe) may solely manifest with epilepsy when segregating to homoplasmy. They may be overlooked in the absence of lactate accumulation and typical mosaic mitochondrial defects in muscle.

Differentiation of morphology, genetics and electric signals in a region of sympatry between sister species of African electric fish (Mormyridae).

Mormyrid fishes produce and sense weak electric organ discharges (EODs) for object detection and communication, and they have been increasingly recognized as useful model organisms for studying signal evolution and speciation. EOD waveform variation can provide important clues to sympatric species boundaries between otherwise similar or morphologically cryptic forms. Endemic to the watersheds of Gabon (Central Africa), Ivindomyrus marchei and Ivindomyrus opdenboschi are morphologically similar to one another. Using morphometric, electrophysiological and molecular characters [cytochrome b sequences and amplified fragment length polymorphism (AFLP) genotypes], we investigated to what extent these nominal mormyrid species have diverged into biological species. Our sampling covered the known distribution of each species with a focus on the Ivindo River, where the two taxa co-occur. An overall pattern of congruence among datasets suggests that I. opdenboschi and I. marchei are mostly distinct. Electric signal analysis showed that EODs of I. opdenboschi tend to have a smaller initial head-positive peak than those of I. marchei, and they often possess a small third waveform peak that is typically absent in EODs of I. marchei. Analysis of sympatric I. opdenboschi and I. marchei populations revealed slight, but significant, genetic partitioning between populations based on AFLP data (F(ST) approximately 0.04). Taken separately, however, none of the characters we evaluated allowed us to discriminate two completely distinct or monophyletic groups. Lack of robust separation on the basis of any single character set may be a consequence of incomplete lineage sorting due to recent ancestry and/or introgressive hybridization. Incongruence between genetic datasets in one individual, which exhibited a mitochondrial haplotype characteristic of I. marchei but nevertheless fell within a genetic cluster of I. opdenboschi based on AFLP genotypes, suggests that a low level of recent hybridization may also be contributing to patterns of character variation in sympatry. Nevertheless, despite less than perfect separability based on any one dataset and inconclusive evidence for complete reproductive isolation between them in the Ivindo River, we find sufficient evidence to support the existence of two distinctive species, I. opdenboschi and I. marchei, even if not 'biological species' in the Mayrian sense.

Severe pneumonia due to Legionella pneumophila: prognostic factors, impact of delayed appropriate antimicrobial therapy.

OBJECTIVE: To compare the outcome of patients with severe Legionella pneumonia (LP) according to the presence or absence of prognostic factors currently reported in the literature and delays in initiating fluoroquinolones and macrolides. DESIGN: Retrospective clinical investigation. SETTING: Intensive care unit (ICU) of an university hospital. PATIENTS: Forty-three consecutive cases with no previous treatment with a macrolide or a fluoroquinolone. MEASUREMENTS AND MAIN RESULTS: The 14 (33%) patients who died of LP were compared with the 29 survivors. Thirty-eight patients (88%) received a fluoroquinolone in combination with a macrolide agent, two patients erythromycin alone and three ofloxacin alone. In univariate analysis, SAPS II more than 46 ( p=0.006) and intubation requirement ( p=0.012) were associated with a higher mortality whereas the administration of fluoroquinolones ( p=0.011) or erythromycin ( p=0.044) within 8 h of arrival on the ICU was associated with better survival. By logistic regression analysis, SAPS II score more than 46 [odds ratio (OR) 8.69; 95% confidence interval (CI) 1.15-66.7; p=0.036], duration of symptoms prior to ICU admission longer than 5 days (OR 7.46; 95% CI 1.17-47.6) were independent risk factors for death. Fluoroquinolone administration within 8 h of ICU arrival (OR 0.16; 95% CI 0.03-0.96; p=0.035) was associated with a reduced mortality. CONCLUSIONS: SAPS II score higher than 46, duration of symptoms prior to ICU admission longer than 5 days and intubation were associated with increased mortality. Initiation of fluoroquinolone therapy within 8 h of ICU admission significantly reduced mortality.

Basal teleosts and the question of elopomorph monophyly. Morphological and molecular approaches.

The methodology used by previous authors to resolve the relationships of the elopomorphan taxa is criticized. The morphological characters that have been proposed to support the monophyly of the Elopomorpha are reviewed and it is shown that most of them are weak. A new hypothesis of relationships is proposed on the basis of nucleotidic sequences of ribosomal RNA 18S, 16S and 12S. In order to really test all the possible relationships, the monophyly of the Elopomorpha was not considered a priori. The tree was rooted on Amia calva and Lepisosteus osseus and the ingroup taxa sampling was subsequently increased. The obtained topology shows that the Elopomorpha is a non-monophyletic taxon. Elopiforms, anguilliforms, albuliforms and notacanthiforms are considered here as four monophyletic, incertae sedis taxa among basal teleosts.

Procalcitonin: a valuable indicator of infection in a medical ICU?

OBJECTIVE: To assess the use of procalcitonin (PCT) for the diagnosis of infection in a medical ICU. DESIGN: Prospective, observational study. PATIENTS: Seventy-seven infected patients and 24 patients with systemic inflammatory response syndrome (SIRS) due to other causes. Seventy-five patients could be classified into sepsis (n = 24), severe sepsis (n = 27) and septic shock (n = 24), and 20 SIRS patients remained free from infection during the study. Plasma PCT and C-reactive protein (CRP) levels were evaluated within 48 h of admission (day 0), at day 2 and day 4. RESULTS: As compared with SIRS, PCT and CRP levels at day 0 were higher in infected patients, regardless of the severity of sepsis (25.2 +/- 54.2 ng/ml vs 4.8 +/- 8.7 ng/ml; 159 +/- 92 mg/l vs 71 +/- 58 mg/l, respectively). At cut-off values of 2 ng/ml (PCT) and 100 mg/l (CRP), sensitivity and specificity were 65% and 70% (PCT), 74% and 74% (CRP). PCT and CRP levels were significantly more elevated in septic shock (38.5 +/- 59.1 ng/ml and 173 +/- 98 mg/l) than in SIRS (3.8 +/- 6.9 ng/ml and 70 +/- 48 mg/l), sepsis (1.3 +/- 2.7 ng/ml and 98 +/- 76 mg/l) and severe sepsis (9.1 +/- 18. 2 ng/ml and 145 +/- 70 mg/l) (all p = 0.005). CRP, but not PCT, levels were more elevated in severe sepsis than in SIRS (p<0.0001). Higher PCT levels in the patients with four dysfunctional organs and higher PCT and CRP levels in nonsurvivors may only reflect the marked inflammatory response to septic shock. CONCLUSION: In this study, PCT and CRP had poor sensitivity and specificity for the diagnosis of infection. PCT did not clearly discriminate SIRS from sepsis or severe sepsis.

Molecular systematics of the African electric fishes (Mormyroidea: teleostei) and a model for the evolution of their electric organs.

We present a new molecular phylogeny for 41 species of African mormyroid electric fishes derived from the 12S, 16S and cytochrome b genes and the nuclear RAG2 gene. From this, we reconstruct the evolution of the complex electric organs of these fishes. Phylogenetic results are generally concordant with earlier preliminary molecular studies of a smaller group of species and with the osteology-based classification of Taverne, which divides the group into the Gymnarchidae and the Mormyridae, with the latter including the subfamilies Petrocephalinae (Petrocephalus) and Mormyrinae (all remaining taxa). However, we find that several genera previously recognized by Taverne are non-monophyletic. Within the Mormyrinae, the genus Myomyrus is the sister group to all the remaining taxa. Other well-supported clades within this group are recovered. A reconstruction of electrocyte evolution on the basis of our best-supported topology suggests that electrocytes with penetrating stalks evolved once early in the history of the mormyrids followed by multiple paedomorphic reversals to electrocytes with non-penetrating stalks.

Phylogenetic relationships of mormyrid electric fishes (Mormyridae; Teleostei) inferred from cytochrome b sequences.

The Mormyridae are African osteoglossomorph freshwater fishes of great interest because of their electric organs. They have become an important model in studies of electrophysiology and behavior but their phylogenetic relationships are poorly known. Phylogenetic relationships among mormyrids were determined by comparing cytochrome b sequences (588 bp) of 27 species belonging to 15 genera. Results showed that the Petrocephalus species (subfamily Petrocephalinae) are the sister-group of all other mormyrids (subfamily Mormyrinae). The monophyly of the Mormyrinae was supported, as well as three original intra-Mormyrinae clades. Three genera, Marcusenius, Pollimyrus, and Brienomyrus, were found to be polyphyletic with high support. Some of these polyphylies are tentatively explained. The results confirmed that the lateral ethmoid bone was lost several times within the Mormyrinae. These findings emphasize the necessity of systematic studies and taxonomic revision of the Mormyridae. The tree obtained from the mitochondrial data showed a single rise of each electrocyte type except for electrocyte with penetrating stalk ("Pa"). Constraining the single occurrence of electrocyte type Pa did not require an excessive number of extra steps (1.86%).

Molecular phylogenetics and the evolution of antarctic notothenioid fishes.

The monophyly of the antarctic fish suborder Notothenioidei and the monophyly of its earliest family the Bovichtidae have been investigated with 12S and 16S mitochondrial DNA sequences. New data from Cottoperca, Pseudaphritis, Harpagifer and several outgroups, in addition to available sequences, show that the bovichtids are paraphyletic. Pseudaphritis is the sister group of all the non-bovichtid notothenioids. The same results are found from two independent genetic markers, the nuclear 28S rDNA and the 12S and 16S mitochondrial rDNA. This reliably refutes a previous hypothesis that placed Pseudaphritis as the sister group of all the remaining notothenioids (including Cottoperca and Bovichtus). Bootstrap analyses show that the Notothenioidei are monophyletic (although members of the suborder Trachinoidei have not been surveyed). Subsequent data from hemoglobin composition confirm the present relationships. After discussions between members of the European Science Foundation (ESF) network during its last two meetings, we point out here some fundamental aspects of comparative biology to improve understanding between the physiologist community and phylogeneticists. The most important points are differences in how the concept of homology is used and differences in the consideration of adaptation. When adaptation is evoked or questioned, endless speculations and untestable scenarios are often developed. We strongly advocate the use of phylogenetic trees for testing hypotheses of adaptation (through multiple character mapping). Such a "research program" in comparative biology has the power to improve knowledge because it can potentially lead to new experiments for testing adaptive hypotheses.

Is the piglet a useful animal model of sudden infant death syndrome?

A prospective study of early death was conducted in a large population of piglets in order to investigate the causes of mortality and determine whether this species could be useful as an animal model of the sudden infant death syndrome (SIDS). 1,921 live-born piglets were closely monitored from birth to 2 months of age. The cause of death was analyzed in all the animals which died during this period. Complete histological, bacteriological and virological examinations of all sudden death animals were compared to identical examinations performed in age-matched control animals. 384 animals (20%) died during the study period and 8 sudden deaths were observed (0.4%). The principal causes of nonsudden death were overlaying (9.4%), hypotrophy (6%), infection (2%) and acute fetal suffering (1.4%). Bacterial infection was found in 6 of the sudden deaths. There was 1 case of suffocation and 1 unexplained sudden death. Compared to controls, there was a significantly greater prevalence of pathology (p < 0.01) and of positive tissue bacteriology (p < 0.05) in sudden death animals. The 6 sudden deaths due to bacterial infections were clearly different from the human syndrome. Suffocation is a known cause of sudden infant death. In the final analysis, only 1 animal (0.5/1000) had an outcome which could be assimilated with SIDS. It is concluded that although there probably exists a syndrome in the piglet equivalent to SIDS, its incidence is very low and major obstacles related to the high level of early mortality in this species hinder investigation.