[Analysis of genotype and phenotype of SEC23B gene in a family affected with congenital dyserythropoietic anemia type II].

OBJECTIVE: To detect potential mutation in a family affected with congenital dyserythropoietic anemia type II (CDA II). METHODS: Targeted sequence capture and next-generation sequencing (NGS) were used to analyze the exons and exon-intron boundaries of the SEC23B gene in a clinically suspected CDA II patient. Genotypes of the relatives were validated by Sanger sequencing. Potential impact of amino acid substitution on the structure and function of SEC23B protein was predicted with MutationTaster and PolyPhen-2. The protein structure was predicted with SWISS-MODEL software. RESULTS: The proband was found to harbor double heterozygous mutations of the SEC23B gene, c.1727T>C (p.F576S) and c.1831C>T (p.R611W), which resulted in amino acid substitutions p.F576S and p.R611W. Both mutations were confirmed by Sanger sequencing. The sister of the proband was found to have carried c.1727T>C (p.F576S), while her father and son have carried c.1831C>T (p.R611W) mutation. In addition, the proband was detected to have carried c.211C>T (p.R71X) of the HFE gene, which resulted in substitution of arginine by a stop codon. The impact of above mutations on the structure or function of protein was predicted to be harmful. Splenectomy and iron chelation therapy have achieved effective improvement of anemia and iron overload. Computer simulation suggested that the mutations have altered the 3D structure of the SEC23B protein. CONCLUSION: The novel compound mutations of c.1727T>C and c.1831C>T of the SEC23B gene probably underlie the CDA II in the family, and there is a strong correlation between the genotype and phenotype.

Efficacy and safety of methylphenidate and ginseng in cancer-related fatigue: a network meta-analysis of randomized controlled trials.

Background: Incidence of cancer-related fatigue (CRF), which can persist 5 to 10 years, is nearly 85% in cancer patients. It severely affects the quality of life and is strongly associated with poor prognosis. As clinical trial data on CRF treated with methylphenidate and ginseng, two potential medicines, has been accumulating, an updated meta-analysis was performed to evaluate and compare the efficacy and safety of the two medicines in CRF. Methods: Randomized controlled trials that investigated methylphenidate or ginseng in the treatment of CRF were identified through a literature search. The primary outcome was CRF relief. Standardized mean difference (SMD) was used to analyze the effect. Results: Eight studies on methylphenidate were included and the pooled SMD was 0.18 [95% confidence interval (95% CI): -0.00 to 0.35, P=0.05]. Five studies on ginseng were included and the SMD was 0.32 (95% CI: 0.17-0.46, P<0.0001). Results of network meta-analysis showed that the order was ginseng, methylphenidate, placebo from high efficacy to low and ginseng was significantly better than methylphenidate (SMD =0.23, 95% CI: 0.01-0.45). Incidences of insomnia and nausea caused by ginseng were significantly lower than those caused by methylphenidate (P<0.05). Conclusions: Both methylphenidate and ginseng can significantly ameliorate CRF. Ginseng may be superior to methylphenidate because ginseng may be more effective and might cause less adverse events. Head-to-head trials with fixed protocol are warranted to identify the optimal medical strategy.

Efficacy of ginseng oral administration and ginseng injections on cancer-related fatigue: A meta-analysis.

BACKGROUND: Up to 90% of patients who are under the active treatment suffer from cancer-related fatigue (CRF). CRF can persist about 10 years after diagnosis and/or treatment. Accumulating reports support that ginseng and ginseng injections are both potential drugs for the treatment of CRF but few studies put them together for analysis. METHODS: Two reviewers independently extracted data in 3 databases (PubMed, Cochrane Library and China National Knowledge Infrastructure) from their inception to May 24, 2021. The primary outcome was the effect of ginseng in alleviating CRF. The secondary outcome was ginseng in alleviating emotional or cognitive fatigue. Standardized mean difference (SMD) was employed. RESULTS: Twelve studies were included to evaluate efficacy of ginseng oral administration and ginseng injections on CRF. The pooled SMD was 0.40 (95% confidence Interval [95% CI] [0.29-0.51], P < .00001). Six studies were included to evaluate efficacy of ginseng oral administration on CRF and the SMD was 0.29 (95% CI [0.15-0.42], P < .0001). The order was 2000 mg/d, 3000 mg/d, 1000 mg/d and placebo from high efficacy to low. Ten studies were included to evaluate efficacy of ginseng injections on CRF and the SMD was 0.74 (95% CI [0.59-0.90], P < .00001). Emotional fatigue was reported in 4 studies, ginseng oral administration in 2 and ginseng injections in 2. The pooled SMD was 0.12 (95% CI [-0.04 to 0.29], P = .15). Cognitive fatigue was reported in 4 studies focusing on ginseng injections and the SMD was 0.72 (95% CI [0.48-0.96], P < .00001). CONCLUSION: Ginseng can improve CRF. Intravenous injection might be better than oral administration. Ginseng injections may alleviate cognitive fatigue. No evidence was found to support that ginseng could alleviate emotional fatigue.

Diagnosis of bronchobiliary fistula by bilirubin crystallization in the alveolar lavage fluid: case reports and literature review.

Bronchobiliary fistula (BBF) refers to the abnormal traffic between the biliary tract and the bronchus. The condition is very rare and usually develops secondary to liver echinococcosis or amebiasis, liver abscess, trauma, biliary obstruction, or tumors. BBF has a high mortality rate and currently, there are no accurate and effective diagnostic methods. This study reports the diagnosis and treatment of two patients with BBF which were confirmed by detecting bilirubin crystallization in the sputum. The first patient was a 45-year-old woman admitted to the hospital with "recurrent cough and lung infection". She had a history of multiple biliary tract surgeries and bilirubin crystallization was detected in bronchoalveolar lavage fluid (BALF) upon examination. Computed tomography (CT) imaging and magnetic resonance cholangiopancreatography (MRCP), together with clinical features, confirmed a diagnosis of BBF. The second patient was a 53-year-old woman admitted to the hospital with coughing and bile-like sputum. She had a history of cholangiocarcinoma surgery and bilirubin crystallization was detected in the cytomorphological BALF examination. Endoscopic retrograde cholangiopancreatography (ERCP) combined with clinical features confirmed a diagnosis of BBF. Both patients recovered after treatment and were discharged from the hospital. The clinical diagnosis of BBF largely relies upon imaging combined with clinical standards, and BALF examinations are rarely performed. This current investigation retrospectively analyzed the diagnosis and treatment of two cases of BBF, and demonstrated that bilirubin crystallization in the BALF may be an important diagnostic indicator for BBF.

Detection of drug resistance-associated genes of multidrug-resistant Acinetobacter baumannii.

The beta-lactamase (BLA) genes, the genes for aminoglycosides-modifying enzymes (AMEs), disinfectant-sulfanilamide resistance (qacEDelta1-sul1) genes, class 1 integrase (intl1) gene, and the qnr gene associated with plasmid-mediated quinolone resistance were analyzed using PCR and verified by DNA sequencing for 31 clinical isolates of multidrug-resistant Acinetobacter baumannii (MDRAB). The organism typing was performed by pulsed-field gel electrophoresis (PFGE). The positive rate of ADC, TEM, PER, and DHA of BLA genes were 100%, 61.3%, 19.4%, and 3.2%, respectively; however, the genes of SHV, OXA-23 group, OXA-24 group, GES, VIM, IMP, and qnr gene were negative. The positive rate of the genes of AMEs for aac (3)-I, aac (6')-I, ant (3")-I, ant (2")-I, aac (3)-II, and aac (6')-II were 67.7%, 45.2%, 29.0%, 22.6%, 12.9%, and 3.2%, respectively. The positive rate of qacEDelta1-sul1 and intl1 were 80.6% and 58.1%, respectively. Six different PFGE clones were found, of which two dominated. The findings show that clinical isolates of MDRAB harbor various kinds of resistance genes.

[Microbiology and clinical analysis of six cases of hospital-acquired pneumonia caused by Acinetobacter baumannii].

OBJECTIVE: To study the microbiology, clinical features and treatment outcomes of hospital-acquired pneumonia (HAP) caused by Acinetobacter baumannii. METHODS: Six cases of HAP, caused by Acinetobacter baumannii producing PER-1 type extended-spectrum beta-lactamase verified by molecular biological methods, were studied in Bethune International Peace Hospital, from January to August 2005. Clinical data were collected and analyzed. Pulsed-field gel electrophoresis (PFGE) were used to analyze the homology of these strains. The genes of beta-lactamase (BLA), aminoglycoside-modifying enzymes (AME), resistant to disinfectant-sulfanilamide (qacEDelta1-sul1), and class 1 integrase (intl1) were analyzed using PCR and verified by DNA sequencing. RESULTS: Among these 6 strains of Acinetobacter baumannii, five were susceptible only to imipenem, and 4 were susceptible to meropenem. The PFGE types identified from these isolates were A1 (n=2) and A2-A5 (n=1, respectively). They all were positive for PER-1, TEM-1, and genes of qacEDelta1-sull and intl1. Three of them harbored genes of aac (3)-I and aac (6')-I. Two had aac (3)-I and ant (3'')-I. One had aac (3)-I. All the patients had severe underlying diseases, and had received mechanical ventilation. They all had received broad spectrum antibiotics within 15 days before Acinetobacter baumannii were identified. Although carbapenem and/or cefoperazone/sulbactam had been used, only 3 patients survived. CONCLUSION: The 6 PER-1 type extended-spectrum beta-lactamase-producing Acinetobacter baumannii isolates were closely related and had complex mechanisms of drug resistance. The prognosis of patients infected by them was poor.

[Whole blood leukocyte phagocytosis assay for Candida albicans based on flow cytometry].

OBJECTIVE: To establish a whole blood leukocyte phagocytosis assay for Candida albicans (C.albicans) based on flow cytometry (FCM). METHODS: C.albicans of mid-logarithmic growth phase was labeled by fluorescence probe carboxyfluorescein diacetate succinimidyl ester (CFDA-SE), and then added into CD45-PC5 pre-stained human whole blood cells at a 10:1 multiplicity of infection (MOI) in 37DegreesCelsius. The cells were incubated for 10, 30 and 60 minutes. Phagocytosis rate of C.albicans by the CD45 positive cells in the blood was determined by FCM. RESULTS: In yeast extract peptone dextrose medium (YPD) and under the conditions of 37DegreesCelsius and 50 mL/L CO2, the logarithmic growth phase of C.albicans SC5314 was from the 5th to 11th hour. C.albicans were well stained by 10 mmol/L CFDA-SE after 30-minute incubation. After 10-, 30- and 60-minute incubation with SC5314 C.albicans with CD45⁺ cells, the phagocytosis rates measured by FCM were (80.1 ± 6.1)%, (83.8 ± 7.7)% and (92.3 ± 11.2)% for the neutrophils, (11.2 ± 3.6)%, (15.8 ± 4.4)% and (27.7 ± 6.8)% for the monocytes and (0.9 ± 0.3)%, (0.8 ± 0.4)% and (5.2 ± 1.6)% for the lymphocytes. CONCLUSION: The method for measuring whole blood leukocyte phagocytosis of C.albicans based on FCM is successfully established, and 30 minutes are the proper incubation time for the phagocytosis assay.

Serological response and diagnostic value of recombinant candida cell wall protein enolase, phosphoglycerate kinase, and ß-glucosidase.

There are no specific signs and symtoms for invasive candidiasis (IC), which makes its diagnosis a challenge. Efforts have been made for decades to establish serological assays for rapid diagnosis of IC, but none of them have found widespread clinical use. Using a systemic candiasis murine model, serological response to recombinant proteins of enolase (rEno1), phosphoglycerate kinase (rPgk1), and ß-glucosidase (rBgl2) were evaluated and rEno1 was found to possess the strongest immunoreactivity, followed by rPgk1 and rBgl2. Likewise, IgG antibody titers to rEno1, rPgk1, and rBgl2 in the positive sera of proven IC patients were determined by ELISA. Results show anti-rEno1 antibody possesses the highest titer, followed by rPgk1 and rBgl2. Antibodies against rEno1, rPgk1, and rBgl2 were detected by ELISA tests in a group of 52 proven IC patients or 50 healthy subjects, The sensitivity, specificity, positive and negative predictive values were 88.5, 90.0, 90.2, and 88.2% for anti-rEno1 detection, 86.5, 92.0, 91.8, and 86.8% for anti-rPgk1 detection, and 80.8, 90.0, 89.4, and 81.8% for anti-rBgl2 detection, respectively. The data clearly demonstrate that the recombinant proteins of Eno1, Pgk1, and Bgl2 are promising candidates for IC serodiagnosis. There's great possibility that the recombinant Eno1 will be more applicable in serodiagnosis and vaccine research on account of its strong serological response.

[Prokaryotic expression and immunogenicity analysis of phosphoglycerate kinase 1 of Candida albicans].

OBJECTIVE: To construct prokaryotic expression plasmids of Candida albicans gene phosphoglycerate kinase 1 (pgk-1) and examine the immunogenicity of the recombinant protein. METHODS: The full-length coding sequence of pgk-1 was amplified by PCR and cloned into the prokaryotic expression vector pET30a. The 6×His-tagged protein was induced by IPTG in E.coli BL-21(DE3) and the recombinant protein was identified by SDS-PAGE and Western blotting. The BALB/c mice were immunized with the purified recombinant protein to evaluate the antigenicity of the recombinant protein by ELISA. RESULTS: The full-length pgk-1 gene was cloned from SC5314 genome and pET-30a-pgk-1 was successfully constructed. The recombinant protein PGK-1 was highly expressed in E.coli with a relative molecule mass of 54 810. ELISA indicated that the titer of the antibody was about 1:1024. CONCLUSION: PGK-1 was successfully expressed by prokaryotic expression system and the recombinant protein showed favorable immunogenicity in mice.

Association of Toll-like receptor 4 gene polymorphism and expression with urinary tract infection types in adults.

BACKGROUND: Innate immunity of which Toll-like receptor (TLR) 4 and CXCR1 are key elements plays a central role in the development of urinary tract infection (UTI). Although the relation between the genetics of TLR4 and CXCR1 and UTI is investigated partly, the polymorphisms and expression of TLR4 and CXCR1 in different types of UTI in adults are not extremely clear. METHODOLOGY/PRINCIPAL FINDINGS: This study investigates the presence of TLR4 A (896) G and CXCR1 G (2608) C polymorphisms in 129 UTI patients using RFLP-PCR. Gene and allelic prevalence were compared with 248 healthy controls. Flow cytometry was used to detect TLR4 and CXCR1 expression in the monocytes of UTI patients and healthy controls. TLR4 (896) AG genotype and TLR4 (896) G allele had higher prevalence in UTI (especially in acute cystitis and urethritis) patients, whereas CXCR1 (2608) GC genotype and CXCR1 (2608) C allele had lower prevalence in UTI patients than controls. TLR4 expression was significantly lower in chronic UTI patients than in acute pyelonephritis or healthy controls. CXCR1 expression was similar in both controls and patients. TLR4 expression in chronic UTI patients after astragalus treatment was higher than pre-treatment. CONCLUSIONS: The results indicate the relationship between the carrier status of TLR4 (896) G alleles and the development of UTI, especially acute cystitis and urethritis, in adults. TLR4 expression levels are correlated with chronic UTI.