[Hearing disorders at the candidates of the National Gendarmerie's competition, Ivory Coast].

Objective: Detect the hearing deficiencies of the candi­da­tes of the gendarmerie's competition. Material and method: Pros­pec­tive study realized over 3 years (2008-2010), in the ENT service of the Gendarmerie to Agban (Abidjan). Examination included an otoscopy and a pure tone audiometry. Results: On 23121 candidates, 1245 had a plug of earwax (5.4%). They were aged from 18 to 25 years old (average age: 22.85 years). Eardrum was pathological in 0.5 % of the cases. Prevalence of hearing loss was 1.5%. Hearing loss included sensorineural hearing loss (56.7%), deafness of transmission (29.4%) and mixed deafness (13.9%). Deafness was unilateral in 79.8% and bilateral in 20.2%. Conclusion: Result of audio­gram will be useful for tracking or assessing cases of noise-induced hearing loss attributable to military service.

[Tympanoplasty: Experience of the ENT department in Yopougon hospital (Ivory Cost)]. / La tympanoplastie: expérience du service ORL du CHU de Yopougon (Côte d'Ivoire).

PURPOSE: Perform an analysis of epidemiological, diagnostic and therapeutic aspects of tympanoplasty in Ivory Coast. MATERIALS AND METHODS: Retro and prospective study over 13 years conducted in the ENT department of Yopougon Teaching Hospital. Our study concerned 104 patients. RESULTS: The study population was dominated by young adults with an average age of 30.61 years. Chronic otitis media accounted for 80.76% of past medical history, a condition that was also the main indication (74.04%). Otoscopy pre- and intraoperative found essentially a non-marginal tympanic perforation (74.03%) and preoperative hearing assessment noted a conductive hearing loss in 72% of cases. The retroauricular surgical incision was performed in all patients. Surgical treatment consisted of a tympanoplasty type I in 76.3% of cases. The anatomical results were good in 81% of cases and functionally, 75% of our patients had interaural intensity difference of less than 20 decibels. CONCLUSION: The tympanoplasty performed in our hospital apparently gives results that are similar to those of western countries to the patients who got a post operative control.

[Sinonasal inverted papilloma: our experiences of diagnostic and surgical care in Côte d'Ivoire]. / Papillome inversé naso-sinusien: notre expérience de la prise en charge diagnostique et chirurgicale en Côte d'Ivoire.

OBJECTIVE: To present our experience of the diagnostic and surgical care of sino-nasal inverted papilloma. MATERIAL AND METHOD: Retrospective study over 14 years (1996-2010) realised in the ENT departments in Côte d'Ivoire. Thirteen inverted papilloma confirmed and operated, after computed tomography, were included (7 men and 6 women, from 8 to 55 years old). RESULTS: The nasal obstruction was the main symptom. Twelve inverted papilloma correspond to Krouse stage T3, and 1 case T4. Surgical removal was performed by lateral rhinotomy (8 cases), Caldwell-Luc (4 cases) and De Lima (1 case). Squamous cell carcinoma was associated in 1 case. Recurrence with malignant transformation was observed in 1 case. CONCLUSION: Surgery by external approach allowed to obtaining satisfactory results. It stays of current events in our countries under equipped. Histopathological examination of the surgical specimen must be compulsory.

[A specific plain X-ray in the diagnosis of elongated styloid apophysis in the Ivory Cost]. / Intérêt du profil oblique menton dégagé dans le diagnostic des apophyses styloïdes allongées en Côte d'Ivoire.

OBJECTIVES: To appreciate the value of a simple radiographic incidence and its effectiveness for the search and the measurement of elongated styloid apophysis (ESA) and to recommend it in the routine search of ESA in practice. PATIENTS AND METHOD: This prospective study was realized over a period of 3 years. It involved the study of ESA from conventional radiographic imaging; the face low incidence in comparison with an experimental incidence, the oblique profile chin lift (OPCL). 101 patients presenting with stylalgia were investigated. RESULTS: Out of the 101 patients, the low face cliché found 49 ESA having an average length of 5 cm. The OPCL allowed to identify 63 ESA, 54 of which were bilateral and 9 unilateral. It also helped identify other associated abnormalities: otitis media (5 cases), dental pathologies (4 cases), dental condensing osteitis (2 cases). The comparative analysis demonstrated a better efficiency of the OPCL in comparison to the standard incidence: the LOW FACE (Roc curve). The OPCL allowed the exposure in one incidence the styloid process and to better appreciate the lower and superior borders of the styloid process giving a better estimate of its measurements. CONCLUSION: It appears from this study that the experimental incidence, the OPCL presents a real interest in the investigation of the ESA but also in the study of the soft tissues of the pharynx, the cervical spine, and the temporo-mastoid region. It also allows the detection of other.

[A case of nasal foreign bodies by tooth migration]. / Un cas de corps étranger des fosses nasales par migration dentaire.

Foreign bodies are a frequent reason for ENT consultation especially in children. Although the nature of the foreign body can be variable, the discovery of a tooth in a nasal cavity is a rare occurrence. The authors report a case of foreign body inside the nasal cavity caused by tooth migration and discuss its pathologic, diagnostic and therapeutic aspects.

The fat-derived hormone adiponectin reverses insulin resistance associated with both lipoatrophy and obesity.

Adiponectin is an adipocyte-derived hormone. Recent genome-wide scans have mapped a susceptibility locus for type 2 diabetes and metabolic syndrome to chromosome 3q27, where the gene encoding adiponectin is located. Here we show that decreased expression of adiponectin correlates with insulin resistance in mouse models of altered insulin sensitivity. Adiponectin decreases insulin resistance by decreasing triglyceride content in muscle and liver in obese mice. This effect results from increased expression of molecules involved in both fatty-acid combustion and energy dissipation in muscle. Moreover, insulin resistance in lipoatrophic mice was completely reversed by the combination of physiological doses of adiponectin and leptin, but only partially by either adiponectin or leptin alone. We conclude that decreased adiponectin is implicated in the development of insulin resistance in mouse models of both obesity and lipoatrophy. These data also indicate that the replenishment of adiponectin might provide a novel treatment modality for insulin resistance and type 2 diabetes.

GRIP1 enhances estrogen receptor alpha-dependent extracellular matrix gene expression in chondrogenic cells.

OBJECTIVE: The role of postmenopause on the pathogenesis of cartilage degeneration has been an open question. We assessed cartilage degeneration in estrogen receptor (ER)alpha null mice and examined the role of glucocorticoid receptor-interacting protein 1 (GRIP1) in the ERalpha-dependent transcription of a type II collagen gene (col2a1) with special reference to a crosstalk with the transforming growth factor (TGF)-beta signaling pathway. METHODS: The vertebral cartilaginous endplate from female ERalpha null mice was subjected to histological analyses. Col2a1 expression of primary chondrocytes (PCs) obtained from ERalpha null mice after 17beta-estradiol (E(2)) and TGF-beta1 stimulation was examined by reverse transcription polymerase chain reaction (RT-PCR). Estrogen response element (ERE) or col2a1 promoter-enhancer luciferase reporter system was used to investigate the crosstalk among ERalpha, GRIP1, and MKK6. Col2a1 expression and glycosaminoglycan (GAG) content were measured in ATDC5 cells treated with GRIP1 small interfering RNA (siRNA). RESULTS: ERalpha deficiency clearly accelerated impairment of the vertebral cartilaginous endplate. E(2) and TGF-beta1 stimulation increased col2a1 expression in PC from wild-type mice, but not that from ERalpha null mice. The same stimulation increased the col2a1 promoter-enhancer reporter activity, and the elevated activity was decreased by dominant-negative ERalpha and p38 mitogen-activated protein kinase (MAPK) inhibitor. GRIP1 increased the E(2)-dependent ERE activation in the presence of ERalpha and constitutive-active MKK6. GRIP1 siRNA repressed col2a1 expression and GAG production in ATDC5 cells. CONCLUSIONS: Crosstalks between ERalpha/GRIP1 and TGF-beta/MKK6/p38 MAPK pathway have protective roles on cartilage metabolism via regulating the extracellular matrices expression. The finding may lead to the development of a novel therapeutic approach for cartilage degeneration.

Assessment of reproductive potential in multiple-spawning fish with indeterminate fecundity: a case study of yellow sea bream Dentex hypselosomus in the East China Sea.

This study examined the spawning season, spawning frequency and batch fecundity of yellow sea bream Dentex hypselosomus in the East China Sea to reassess the previously reported reproductive characteristics of the species. Time-course sampling showed that this species had a diurnal ovarian maturation rhythm. Late tertiary yolk-stage oocytes appeared 2 days before spawning, starting the process of germinal vesicle movement and breakdown. On the day of spawning, ovulation and subsequent spawning occurred in the early morning (0400-0800 hours). Postovulatory follicles disappeared from the ovaries within c. 24 h of ovulation. Seasonal changes in the ovarian conditions indicated that this species spawned more or less throughout the year, with the peak ranging from spring to autumn. The compositions of the developing oocytes and degenerating postovulatory follicles in the ovaries suggested that most females spawned repeatedly over 2 to 3 consecutive days during the peak of the spawning season. Somatic body condition did not have a significant effect on batch fecundity, but there was a significant relationship between batch fecundity and fork length according to spawning status. Females spawning on consecutive days were more fecund than those spawning every other day. The findings show that this species has much greater reproductive potential than previously estimated.

Effect of Divalent Ions on Electroosmotic Flow in Microchannels.

The electroosmotic flow (EOF) rate in fused silica microchannels is experimentally found to decrease when trace quantities of salts containing the divalent cations Ca(2+) and Mg(2+) are added to a constant ionic strength background electrolytic solution (BGE) flowing in a channel having negatively charged walls. Moreover, the observed effect is quantitatively different for the two ions Ca(2+) and Mg(2+). Since electrostatic interactions are identical for ions of the same valence modeled as point charges, a description of the electric double layer (EDL) based on the Poisson-Boltzmann equation alone cannot account for these experimental observations. New experimental observations on electroosmotic flow in presence of Ca(2+) and Mg(2+) are reported in this work. A site binding model that accounts for the chemical interactions of the BGE ions with the silica surface is developed. The model predictions are in good agreement with the experimental observations on divalent cations as well as data from the literature on how properties such as pH and ionic strength affect electroosmotic flow rates for a BGE with monovalent cations.

Cardiac retransplantation: a 15-year single-center clinical experience.

BACKGROUND: Cardiac retransplantation is a controversial therapy because of the shortage of donor hearts. We retrospectively reviewed the short-term and long-term outcomes after cardiac retransplantation. METHODS AND RESULTS: Twenty-eight cases (18 males, 7 females; mean age, 50.3 +/- 13.5 years) underwent cardiac retransplantation: 25 first retransplantations and 3 second retransplantations. The indications for retransplantation were primary graft failure (PGF) in 11 patients (39.3%), intractable acute cardiac rejection (IACR) in 4 patients (14.3%), and coronary allograft vasculopathy (CAV) in 13 patients (46.4%). The patients had been supported as follows: prolonged cardiopulmonary bypass (CPB; n = 3), intra-aortic balloon pumping (IABP; n = 1), intravenous inotropic support (n = 7), extracorporeal membranoxygenator (ECMO; n = 3), ventricular assist device (VAD; n = 4), and no inotropic support (n = 10). There were 8 deaths within 30 days after retransplantation (28.6%). The overall 1-, 5-, 10-, and 15-year survival rates were 46.4%, 40.6%, 32.5%, and 32.5%, respectively. Acute cardiac rejection was the most common cause of death (43.8%). Thirty-day and 1-year survival rates of IACR, PGF, and CAV were 50.0%/0%, 63.6%/45.5%, and 84.6%/68.4%, respectively. CONCLUSIONS: Long-term survival after retransplantation was acceptable for patients with CAV and PGF; however, we must select patients for retransplantation carefully if the indication is IACR, because of the poor outcome.

Particularities of bacterial ecology of acute otitis media in an African subtropical country (Cote d'Ivoire).

BACKGROUND: Acute otitis media (AOM) is the commonest infectious disease of childhood. Its treatment is based on a probabilistic antibiotic therapy. OBJECTIVE: To determine the prevalence of different bacteria responsible for AOM in children in Abidjan (Côte d'Ivoire). MATERIALS AND METHODS: This is a prospective study, between November 2002 and August 2003. To identify bacterial isolates, 60 children under 15 years old with AOM were enrolled. In 52 cases, the sample was recovered by aspiration of a spontaneous otorrhea and in eight cases, it was taken by paracentesis. Nasopharyngeal swabs were cultured. Antibiotic susceptibility tests were performed on isolates. RESULTS: 75% of children were under 24 months of age. Pseudomonas aeruginosa was recovered in 31.8% of cases followed by Streptococcus pneumoniae (21%), Proteus (7.4%) and Staphylococcus (6.2%). The rate of penicillin-nonsusceptible S. pneumoniae (NSSP) was 11.8%. Resistance to erythromycin was 6%. For Pseudomonas, resistances to antibiotics were 35.5% to ceftazidim and 16% to pefloxacin, respectively, but all the strains were sensible to colistin, iminepem and ciprofloxacin. CONCLUSION: These data can modify therapeutic approaches for treatment of AOM in our country. This study should encourage surveillance of AOM microbiology.

Study on the introduction of hazard analysis and critical control point (HACCP) concept of the water quality management in water supply systems.

In the latest revision in 2004, the 3rd edition, the Water Safety Plans (WSP) was newly introduced into the World Health Organization (WHO) Guidelines for Drinking Water Quality. The Hazard analysis and critical control point (HACCP) is a basic concept that underlies the WSPs, and is also known as the product quality management method in the field of food and the medical manufacturing industries. In the amendments of the Drinking Water Quality Standards in Japan, water suppliers are required to reasonably achieve both safe water and efficient water quality management. Therefore, the HACCP concept is focused as an adequate management method covering a whole process of water supply systems, in a systematic way. The purpose of this study is to investigate a practical procedure in introducing the HACCP into water quality management in Japan. In comparison to conventional applications of the HACCP, unmanageable variations of raw water quality, continuous treatment and supply, and numerous standards of water quality items need to be considered. The HACCP system is expected to achieve a quick response to improvements in water quality, accountability towards consumers and a decrease in accidents.

Anterolateral right thoracotomy for mitral valve procedure after previous coronary artery bypass grafting with functioning internal mammary artery grafts.

AIM: Mitral valve procedure after previous coronary artery bypass grafting (CABG) with functioning internal mammary artery (IMA) grafts has high risk. Especially, internal mammary artery grafts injury may be fatal. The anterolateral right thoracotomy affords easy access to the right atrium with minimal dissection, and minimizes the risk of injury to the IMA grafts. We reviewed our operative technique and outcome after mitral valve procedure after previous CABG with functioning IMA grafts. METHODS: Thirteen patients (11 male and 2 female, mean age of 67.7+/-8.5 years, range 54 to 80 years) underwent mitral valve replacement after previous CABG with functioning IMA grafts from march 1993 to september 2002. The mean interval between the previous CABG and the mitral valve procedure was 3.8 years (range 9 months to 8 years). Four patients had simultaneous mitral valve procedures at initial CABG (2 repairs and 2 replacements). The operation has performed through the anterolateral right thoracotomy, under ventricular fibrillation with moderate hypothermia and without cardioplesia. RESULTS: Mitral valve repair was performed in 3 patients, mitral valve replacement in 10 patients. The mean coronary bypass time was 69.1+/-16.2 min (range 45 to 98 min). The operation time was 159.3+/-29.4 min (range 120 to 219 min). Intensive care unit stay days was 1.9+/-1.6 days (range 1 to 5 days). Peak CK/CK-MB values were 555.1+/-290.4 IU/16.6+/-10.7 IU (range 176 to 924 IU/7 to 44 IU). Peak troponin I value was 9.5+/-5.2 pg/mL (range 4 to 17.8 pg/mL). There was no IMA injury and no early death. Other complications were newly arrhythmia in 3 patients, renal insufficiency in 1 patient, reoperation for bleeding in 1 patient. CONCLUSIONS: Anterolateral right thoracotomy approach, ventricular fibrillation with moderate hypothermia without cardioplesia were a safe and good method for mitral valve operation after previous CABG with functioning IMA graft.

Macrophage colony-stimulating factor induces interleukin-8 production in human monocytes.

We have investigated the stimulatory effect of recombinant human macrophage colony-stimulating factor (rhM-CSF) on interleukin-8 (IL-8) production by human peripheral blood monocytes. When monocytes were prepared from peripheral blood and cultured for 24 hours, the average amount of IL-8 in the culture medium was about 8.9 +/- 3.2 ng/2 x 10(5) cells. In contrast, the production of IL-8 by monocytes increased to a level of 19.2 +/- 4.7 ng/2 x 10(5) cells in response to rhM-CSF. This induction by rhM-CSF was dose-dependent. Northern blot analysis showed that expression of IL-8 at the pretranslational level was enhanced after M-CSF treatment. Kinetic studies showed that secretion of IL-8 from monocytes was enhanced within 2 hours after exposure to rhM-CSF, and a saturation level, which was reached around 48 hours, was two-fold higher than that of cells without M-CSF treatment. In addition, conditioned medium of M-CSF-stimulated monocytes activated the chemotaxis of human neutrophils, and this activity was significantly inhibited by anti-IL-8 antibody. These results, taken together, suggest that M-CSF can affect many cellular functions through regulation of IL-8 expression in monocytes.

[Epidemiology and treatment of sinus mucoceles: 25 cases in Abidjan (Côte d'Ivoire)]. / Les mucoceles sinusiennes: etude epidemio-clinique et therapeutique de 25 cas colliges a Abidjan (Cote d'Ivoire).

OBJECTIVE: To determine the epidemiological, clinical and therapeutic characteristics of sinus mucoceles in Ivory Coast. MATERIALS AND METHODS: A multicenter study conducted in the three services ENT (ear, nose & throat) hospital and university centers and clinics in Abidjan from 1996 to 2012 (16 years). It involved patients who have been diagnosed clinically and radiologically with a sinus mucocele who benefited from surgical treatment. RESULTS: During the study period, 25 patients were recruited - a hospital incidence of 1.5 cases per year. A female predominance was noted with 60% of women and 40% men with a sex ratio of 0.67. Patients consulted for headache (68%), facial swelling (68%), proptosis (28%). Allergic rhinitis is the antecedent rhinologic found most commonly (32%) followed by sinusitis (12%) and nasal sinus surgery (12%). The naso-sinus CT was performed in all patients diagnosed with mucocele. Surgical excision was made by several surgical approach, the most used were the way of JACQUES and paralateronasal rhinotomy of MOURE. Pathological examination of the surgical specimen confirmed the diagnosis of sinus mucocele. The development was beset by complications such as eyelid swelling (1 case), blurred vision (1 case), unsightly surgical scar (1 case). CONCLUSION: The nasal sinus mucoceles are relatively rare benign lesions with pseudo-cystic insidious evolution. Diagnosis suspected by the clinic is confirmed by radiology. Their treatment is mainly surgical.

OBJECTIF: Déterminer les caractéristiques épidémio-cliniques et thérapeutiques des mucocèles sinusien-nes en Côte d'Ivoire. MATÉRIEL ET MÉTHODE: Il s'est agit d'une étude multicentrique menée dans les services d'ORL des trois centres hospitaliers et universitaires et les polycliniques d'Abidjan de 1996 à 2012 (16 ans). Elle a concerné les patients chez qui il a été diagnostiqué cliniquement et radiologiquement une mucocèle sinusienne et qui ont bénéficié d'une prise en charge chirurgicale. RÉSULTATS: Pendant la période d'étude, 25 patients ont été recrutés soit une incidence hospitalière de 1.5 cas par an. Il y avait une prédominance féminine avec 60% de femme et 40% d'homme soit un sex ratio de 0.67. Les patients consultaient pour des céphalées (68%), des tuméfactions faciales (68%), une exophtalmie (28%). La rhinite allergique est l'antécédent rhinologique le plus retrouvé (32%) suivi des sinusites (12%) et de la chirurgie naso-sinusienne (12%). La tomodensitométrie naso-sinusienne réalisée chez tous nos patients, a posé le diagnostic de mucocèle dans 100% des cas. L'exérèse chirurgicale a été faite par plusieurs voie d'abord dont les plus utilisées étaient la voie de JACQUES et la rhinotomie paralatéronasale de MOURE. L'examen anatomopathologique des pièces opératoires a confirmé le diagnostic de mucocèle sinusienne. L'évolution a été émaillée de complications à type de tuméfaction palpébrale (1 cas), flou visuel (1 cas), cicatrice opératoire disgracieuse (1 cas). CONCLUSION: Les mucocèles naso-sinusiennes sont des lésions bénignes pseudo-kystiques relativement rares, d'évolution insidieuse. Leur diagnostic soupçonné par la clinique est confirmé par le bilan radiologique. Leur traitement est essentiellement chirurgical.

Organization of the gene for gelatin-binding protein (GBP28).

GBP28 is a novel human plasma gelatin-binding protein that is encoded by apM1 mRNA, expressed specifically in adipose tissue. Three overlapping clones (two lambda clones and one BAC clone) containing the human plasma gelatin-binding protein (GBP28) gene were isolated and characterized. The GBP28 gene spans 16kb and is composed of three exons from 18bp to 4277bp in size with consensus splice sites. The sizes of the two introns were 0.8 and 12kb, respectively. The gene's regulatory sequences contain putative promoter elements, but no typical TATA box. The third exon of this gene contains a long 3'-untranslated sequence containing three Alu repeats. The exon-intron organization of this gene was very similar to that of obese gene, encoding leptin. We also report the chromosome mapping of this gene by fluorescence in situ hybridization (FISH) using a genomic DNA fragment as a probe. The GBP28 gene was located on human chromosome 3q27. The nucleotide sequence data reported in this paper will appear in the DDBJ/EMBL/GenBank nucleotide sequence databases with the accession numbers ABO12163, ABO12164 or ABO12165.

Bond strength of binary titanium alloys to porcelain.

The purpose of this study was to investigate the bond strength between porcelain and experimental cast titanium alloys. Eleven binary titanium alloys were examined: Ti-Cr (15, 20, 25 wt%), Ti-Pd (15, 20, 25 wt%), Ti-Ag (10, 15, 20 wt%), and Ti-Cu (5, 10 wt%). As controls, the bond strengths for commercially pure titanium (KS-50, Kobelco, Japan) and a high noble gold alloy (KIK, Ishifuku, Japan) were also examined. Castings were made using a centrifugal casting unit (Ticast Super R, Selec Co., Japan). Commercial porcelain for titanium (TITAN, Noritake, Japan) was applied to cast specimens. The bond strengths were evaluated using a three-point bend test according to ISO 9693. Since the elastic modulus value is needed to evaluate the bond strength, the modulus was measured for each alloy using a three-point bend test. Results were analyzed using one-way ANOVA/S-N-K test (alpha = 0.05). Although the elastic moduli of the Ti-Pd alloys were significantly lower than those of other alloys (p = 0.0001), there was a significant difference in bond strength only between the Ti-25Pd and Ti-15Ag alloys (p = 0.009). The strengths determined for all the experimental alloys ranged from 29.4 to 37.2MPa, which are above the minimum value required by the ISO specification (25 MPa).

Induction of apoptosis in various tumor cell lines by geranylgeraniol.

We have found that GGO is a potent inducer of apoptosis in various human tumor cell lines, including a myeloid multipotential leukemia K562 cell line which is resistant to the induction of apoptosis by various apoptosis-inducing agents and conditions. Polyprenylalcohols with isoprenyl units shorter than the geranylgeranyl group or longer than farnesyl group were less effective in inducing apoptosis in myeloid leukemia HL-60 cells and polyprenylketones had no apoptosis-inducing activity. The fragmentation of DNA in HL-60 cells by GGO was so rapid, no significant effect on the cell cycle was observed. [3H]GGO was taken up by HL-60 cells in 3 h and was incorporated into several proteins. The expression of c-myc and bcl-2 decreased significantly within 3 h of the start of the treatment of HL-60 cells with 50 microM GGO.

Serial neuroimages of acute necrotizing encephalopathy associated with human herpesvirus 6 infection.

A previously healthy 8-month-old girl developed exanthem subitum and acute encephalopathy with status epilepticus, quadriplegia and bilateral abducens nerve palsies. Human herpesvirus-6 DNA was found in the cerebrospinal fluid by the polymerase chain reaction at the acute stage. Cranial computed tomography showed low density areas in the thalami and in the cerebellar and abducens nuclei. The distribution of the lesions was consistent with acute necrotizing encephalopathy. As for the thalamic lesions, a T2 weighted magnetic resonance image on the 24th day of the illness demonstrated low signal intensity surrounded by high intensity; 99mTc-ECD SPECT showed hypoperfusion, which suggested irreversible tissue damage. The patient is now 1 year 6 months old and has spastic quadriparesis with mental retardation and abducens nerve palsies.

Cloning and sequencing of the chromosomal DNA and cDNA encoding the mitochondrial citrate synthase of Aspergillus niger WU-2223L.

The complementary DNA (cDNA) and chromosomal DNA encoding the citrate synthase (EC 4.1.3.7) gene (cit1) of Aspergillus niger WU-2223L, a citric acid-producing strain, were cloned. Synthetic oligonucleotide primers were designed according to the amino acid sequences of already known eukaryotic citrate synthases and the codon bias of A. niger genes. The 920-bp DNA fragment was amplified by polymerase chain reaction with these primers using chromosomal DNA of WU-2223L as a template, and was employed to screen a cDNA library of A. niger. One full-length cDNA clone was isolated and sequenced, within which an ORF of 1425 by encoding a protein of 475 as with a molecular weight of 52,153 Da was found. Its N-terminal region contains a typical mitochondrial-targeting motif. The predicted as sequence was 82, 68, and 65% homologous with the mitochondrial citrate synthases of Neurospora crassa, Saccharomyces cerevisiae, and pig, respectively, but it showed lower homology to bacterial citrate synthases. The full-length cDNA clone was used to screen a chromosomal library of A. niger WU-2223L, and a 7.5 kb-SalI fragment containing the corresponding chromosomal gene was isolated. Comparison of the chromosomal and cDNA sequences revealed that the cit1 gene is interrupted by six introns. In the chromosomal DNA, upstream of the coding region, a CT-rich region, but not the TATAAA or CAAT motifs, was found. Escherichia coli MOB150, a citrate synthase-deficient mutant showing a glutamate-requiring phenotype, was transformed with the plasmid pKAC-35S, which is the expression vector pKK223-3 containing the cDNA fragment encoding a putative mature protein of A. niger citrate synthase. The transformant harboring pKAC-35S showed citrate synthase activity and a glutamate-nonrequiring phenotype.