Drug testing involves many analytical instruments and test items, sample pretreatment is tedious, the industry's intelligence level remains low, making drug testing a labour-intensive job. However, in the era of Industry 4.0 intelligent manufacturing, intelligent transformation of the quality control (QC) laboratory has become the focus of industry. At the same time, driven by consistency evaluation of the quality and efficacy of generic drugs and the centralized procurement policies, pharmaceutical companies have intensified their competition, further stimulating the intrinsic demand for laboratory intelligence. Based on the current state and future trends of the pharmaceutical industry, this review discusses the development of a digital and automated QC laboratory. It points out the necessity of transitioning from the traditional centralized laboratory model to an intelligent, distributed quality control model to accommodate continuous manufacturing processes. At the same time, it also analyses the potential challenges in the implementation process and coping strategies, in order to provide relevant practitioners with ideas for building intelligent QC laboratories.

Abstract: Objective　To identify the species of Mycobacteroides abscessus complex (MABC) in patients with pulmonary infection from the Second Affiliated Hospital of Hainan Medical University, and to investigate the species types, drug sensitivity and population distribution of MABC in pulmonary infection in Hainan. Methods　Respiratory tract specimens were collected from suspected tuberculosis patients who visited the Second Affiliated Hospital of Hainan Medical University from January 2014 to December 2021 and cultured for Mycobacterium isolation. Non-tuberculous mycobacteria (NTM) strains were preliminarily identified by p-nitrobenzoic acid/thiophen-2-carbohydrazide (PNB/TCH) medium and DNA microarray chip, and then MABC and its subspecies were identified by hsp65 and rpoB gene sequencing. In vitro antimicrobial susceptibility test was performed by broth microdilution method. Results　A total of 3 025 respiratory specimens from suspected pulmonary tuberculosis patients were collected during the study period. Among the 123 patients with identified MABC isolates, 124 MABC strains were isolated and identified, including 74 strains of Mycobacteroides abscessus subsp. abscessus, 38 strains of Mycobacteroides abscessus subsp. massiliense and 12 strains of Mycobacteroides abscessus subsp. bolletii. Among them, 118 patients had single MABC subspecies infection, one patient had mixed infection with two MABC subspecies, two patients had mixed infection with MABC and other NTM, and two cases had mixed infection with MABC and M.tuberculosis. There were more female patients than male patients with a ratio of 1:0.64, and those aged 50 and above amounted to 76.42% (94/123, 95%CI: 67.93%-83.61%). There was no significant difference in age distribution between male and female patients (Z=-0.944, P=0.347). The drug susceptibility results showed that all MABC strains were sensitive to Tigecycline (TGC), with a resistance rate of 0.81% (1/124) to Amikacin (AK), and resistance rates of 6.45% (8/124), 32.26% (40/124), and 74.19% (92/124) to Cefoxitin (FOX), Linezolid (LZD), and Imipenem (IPM), respectively. For Clarithromycin (CLR), MABC showed induced resistance , and there was a statistically significant difference in the CLR (14D) resistance rates among the three subspecies (χ2=66.335, P<0.001). The resistance rates to Tobramycin (TOB), Doxycycline (DOX), Moxifloxacin (MFX), Ciprofoxacin (CIP), Trimethoprim/Sulfamethoxazole (TMP-SMX), and Amoxicillin/Clavulanic acid (AMC) were high, all >80%. Conclusion　 In Hainan Province, pulmonary infections with MABC are mainly caused by Mycobacteroides abscessus subsp. Abscessus, which show high rates of inducible resistance to CLR. Timely and accurate identification of MABC to subspecies and drug susceptibility testing are of significant important for clinical decision-making.

OBJECTIVETo research the animal model with mimetic aging effect in the inner ear predispose to the ototoxicity of kanamycin.

METHODSFifty wistar rats were randomly divided into four groups: group A (D-galactose group, n = 14) were treated with hypodermic 5% D-galactose (150 mg x kg(-1) x d(-1)) for 8 weeks and then with intraperitoneal saline for 10 days; group B (D-galactose and kanamycin group, n = 14) were given the same dose of D-galactose but kanamycin (500 mg x kg(-1) x d(-1)) instead of saline; group C (kanamycin group, n = 12) were treated with saline for 8 weeks and then with intraperitoneal kanamycin for 10 days;group D (control group, n = 10) were given saline only. Auditory brainstem response (ABR) was used to detect the hearing threshold of rats and colorimetry was used to analyze the activity of the GSH-PX. The inner ear tissue was harvested and the mitochondrial DNA was amplified to identify the 4834 bp deletion mutation by nested primer polymerase chain reaction (nested PCR) technique.

RESULTSThe incidence of mitochondrial DNA 4834 bp deletion mutation was 100% (28/28) in group A, 92.86% (26/28) in group B and 0% in group C or group D. The activity of GSHPX in group A was (59.07 +/- 8.70)U, (63.29 +/- 12. 40)U in group B, (136.67 +/- 9.53)U in group C and (142.10 +/- 7.02)U in group D. The difference between group A and D was significant (P = 0.000) while the difference between group A and B was not significant (P = 0.307), which was similarly as between group C and group D (P = 0.151). ABR threshold was (5.36 +/- 3.08) dB peSPL in group A, (61.79 +/- 11.20) dB peSPL in group B, (34.17 +/- 4.69) dB peSPL in group C and (6.50 +/- 3.37) dB peSPL in group D. No difference was found between group A and D (P = 0.398) while the difference in shift of ABR threshold between group B and group C (or group D) was significant (P = 0.000).

CONCLUSIONSThe mimetic aging effect in the inner ear of the rat can be induced by D-galactose, and these rats present high incidence of mtDNA4834 deletion which can greatly enhance the sensitivity of the inner ear to the kanamycin.

Aging, Premature ; chemically induced ; Animals ; DNA, Mitochondrial ; genetics ; Disease Models, Animal ; Disease Susceptibility ; Ear, Inner ; drug effects ; Galactose ; toxicity ; Kanamycin ; toxicity ; Rats ; Rats, Wistar ; Sequence Deletion

BACKGROUNDMitochondrial DNA mutations have been found in sensorineural deafness. The aim of this study was to compare three methods for extraction of nucleic acid from membranate inner ear tissue of rats.

METHODSAlkaline denaturation, a conventional phenol-chloroform method and Trizol reagent were respectively used to extract the slight nucleic acid from membranate inner ear tissue of rats. We assessed the amount and quality of nucleic acid using a UV-spectrometer and polymerase chain reaction (PCR).

RESULTSThe yield and purity (OD260/OD280) of DNA from inner ear tissue using the phenol-chloroform method was the highest of the three methods. Mitochondrial DNA (mtDNA) fragment can be amplified by PCR from nucleic acid prepared by all methods, while no nuclear DNA (nDNA) fragment can be amplified by method of alkaline denaturation. Both nuclear and mitochondrial genes could be amplified by reverse transcriptional PCR from the RNA prepared by Trizol reagent.

CONCLUSIONAdequate amount and high-quality of mtDNA, nDNA and RNA were obtained from unilateral membranate inner ear tissue of rats. Method of alkaline denaturation could be chosen when mtDNA without nDNA was needed, while phenol-chloroform method was suitable for extracting total DNA (including nDNA and mtDNA); method with Trizol reagent was suitable for extracting total RNA and total DNA.

Animals ; DNA ; isolation & purification ; DNA, Mitochondrial ; isolation & purification ; Ear, Inner ; chemistry ; Polymerase Chain Reaction ; RNA ; isolation & purification ; Rats ; Rats, Wistar

OBJECTIVETo investigate the application of fluorescence in situ hybridization (FISH) technique in prenatal diagnosis of complex chromosomal abnormalities.

METHODSEleven prenatal diagnosis cases (8 from amniocentesis and 3 from cord blood) with complex chromosomal abnormalities detected by routine G-banding, were further analyzed by FISH.

RESULTSThe FISH technique confirmed the results of balanced chromosome rearrangements detected by G-banding, and clarified the structure of the derivative chromosomes in the 3 amniocentesis samples and the origin of the mark chromosomes in the 2 cord blood samples.

CONCLUSIONFISH can be used to diagnose the complex chromosomal abnormalities accurately in prenatal diagnosis, and can provide very useful genetic information for clinical diagnosis and treatment.

Amniotic Fluid ; chemistry ; Chromosome Aberrations ; Female ; Fetal Blood ; chemistry ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Pregnancy ; genetics ; Prenatal Diagnosis ; methods

Leigh syndrome is a genetically heterogeneous disease caused by defects in enzymes involved in aerobic energy metabolism and the Krebs', cycle. Mitonchondrial complex I deficiency is a main cause of Leigh syndrome. In this study, a Chinese child with Leigh syndrome caused by 13513G>A mutation was reported. The proband was the first child of his parents. The previously healthy boy presented with generalized epilepsy at 12 years of age. When he visited Peking University First Hospital at 13 years of age, he had subacute loss of vision in two eyes and temporal defect of visual field in the left eye. He walked with a spastic gait. His blood lactate and pyruvate levels were elevated. Muscle biopsy showed mild lipid accumulation in muscle fiber. An electrocardiogram showed incomplete right bundle branch block. Brain magnetic resonance imaging showed bilateral, symmetrical lesions in the basal ganglia, supporting the diagnosis of Leigh syndrome. 13513G>A mutation was identified by gene analysis in the patient, which led to mitochondrial respiratory chain complex I deficiency. Multivitamins and L-carnitine were administered. At present, the patient is 16 years old and has progressive deterioration with significant muscle weakness and body weight loss. He is absent from school. He has no obvious retardation in intelligence. 13513G>A mutation was first identified by gene analysis in Chinese population with Leigh syndrome. This may be helpful in genetic counseling.
Adolescent ; DNA, Mitochondrial ; genetics ; Electron Transport Complex I ; deficiency ; Humans ; Leigh Disease ; genetics ; Male ; Mutation

OBJECTIVETo assess the value of fluorescence in situ hybridization (FISH) and bacterial artificial chromosome FISH (BAC-FISH) for the diagnosis for patients with marker chromosomes.

METHODSSixteen patients with marker chromosomes were analyzed with technologies including GTG-banding, Q-banding, multiplex FISH and BAC-FISH.

RESULTSThe marker chromosomes in the 16 patients were verified as der(Y) (2 cases), psu dic(Y) (1 case), psu dic(15) (1 case), dic(15) (1 case), del(Y) (1 case), r(X) (5 cases), i(14 or 22) (2 cases), i(18) (1 case).

CONCLUSIONFISH and BAC-FISH can both verify the origin of marker chromosomes and provide accurate information for the diagnosis and treatment of patients.

Adolescent ; Adult ; Child ; Chromosome Aberrations ; Female ; Genetic Diseases, Inborn ; diagnosis ; genetics ; Genetic Markers ; genetics ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Male ; Young Adult

This study reviews a case of mitochondrial respiratory chain complex I deficiency due to the 10191T>C mutation in mitochondrial ND3 gene. The previously healthy boy progressively presented with blepharoptosis, weakness, epilepsy and motor regression at age 6 years. Elevated blood lactate and pyruvate were observed. Brain magnetic resonance imaging showed symmetrical lesions in the basal ganglia. Leigh syndrome was thus confirmed. The protein from the mitochondria and genomic DNA of the boy and his parents was collected from peripheral blood leucocytes for the activity test for mitochondrial complex I to V and genetic analysis. The results showed the activity of complex I (33.1 nmol /min in 1 milligram mitochondrial protein) was lower than normal reference value (44.0±5.4 nmol /min in 1 milligram mitochondrial protein). The ratio of complex I to citrate synthase (19.8%) was also lower than normal reference value (48%±11%). The activities of complexes II to V were normal. 10191T>C mutation in ND3 gene of mitochondria was identified in the boy. 10191T>C mutation and complex I deficiency were not detected in his parents. At present, he is 16 years old, and of normal intelligence with spastic paralysis in both lower extremities after treatment. It is concluded that a Chinese boy with isolated complex I deficiency due to 10191T>C mutation in ND3 gene was firstly diagnosed by peripheral leukocytes mitochondrial respiratory chain enzyme assay and gene analysis. This study can provide clinical data for the nosogenesis of Leigh syndrome.
Adolescent ; Brain ; pathology ; Electron Transport Complex I ; deficiency ; genetics ; Humans ; Leigh Disease ; genetics ; Magnetic Resonance Imaging ; Male ; Mitochondrial Diseases ; genetics ; Mutation

An ultra performance liquid chromatography-tandem mass spectrometric (UPLC-MS/MS) method was developed for the determination of 11 kinds of aminoglycosides (AGs), including paromomycin, spectinomycin, tobramycin, gentamycin, kanamycin, hygromycin B, apramycin, streptomycin, dihydrostreptomycin,amikacin and neomycin in aquatic products. Samples were extracted by phosphate buffer solution, and purified on molecularly imprinted polymers (MIP) solid phase extraction column. After separated by Obelisc R chromatographic column, AGs were detected by UPLC-MS/MS. It showed a good linearity relationship in the AGs concentration range of 1.0-1000 ng/mL with the correlation coefficient R2>0.994. The limit of detection (LOD,S/N≥3) was ranged from 1.0 μg/kg to 10.0 μg/kg,and the limit of quantitation (LOQ,S/N≥10) was ranged from 2.0 μg/kg to 20.0 μg/kg. Besides, the average recoveries presented 78.4%-109.6% with the relative standard deviation (RSD, n=6) of 2.3%-14.9%. This method was successfully applied to the simultaneous determination of 11 kinds of AGs with high sensitivity in aquatic products.

OBJECTIVE To investigate the antimicrobial resistance of hospital-and community-acquired pathogens collected from 10 teaching hospitals located at different areas in China in 2006.METHODS According to the study protocol,the strains of Streptococcus pneumoniae,meticillin-susceptible Staphylococcus aureus(MSSA),Escherichia coli and Klebsiella pneumoniae were collected and sent to the central lab for reidentification and susceptibility testing.The minimal inhibitory concentrations(MICs) of antimicrobial agents against Str.pneumoniae were determined by Etest method and MICs of antimicrobial agents against S.aureus,E.coli and K.pneumoniae strains were determined by agar dilution method.WHONET5.4 software was used to analyze the data.RESULTS Among 353 Str.pneumoniae strains,74.2% were penicillin-susceptible(PSSP),9.6% were penicillin-intermediate(PISP) and 16.2% were penicillin-resistant(PRSP).Strains from different hospitals showed different sensitivity to penicillin.Among ?-lactam antibiotics,cefuroxime showed the lowest susceptibility rate of 0%(for PRSP) to 76.7%(for PSSP).The susceptibility rate to ceftriaxone and amoxicillin-clavulanic acid was 98.1% and 98.9% in PSSP group,61.8% and 64.7% in PISP group,and 15.8% and 10.5% in PRSP group.The ESBLs rate was 56.2% among 267 Escherichia strains and 42.7% among 206 K.pneumoniae strains.For ESBLs-producing strains,the susceptibility rates to cefotaxime and ceftriaxone were low and the rate to ceftazidime was relatively high among ?-lactam antibiotics.73.4% MSSA strains produced ?-lactamase.?-Lactam antibiotics tested showed high susceptibility against MSSA strains.The susceptibility rate was 98.9-100%.The susceptibility rate to ciprofloxacin and levofloxacin was 80.8% and 88.1%,separately.CONCLUSIONS Fluoroquinolones show high susceptibility against Str.pneumoniae.Ceftriaxone and amoxicillin-clavulanic acid have relatively high susceptibility among ?-lactams.For MSSA and non-ESBLs-producing E.coli and K.pneumoniae strains,?-lactams show high susceptibility.For ESBLs-producing E.coli and K.pneumoniae strains,the susceptibility rates to cefotaxime and ceftriaxone are low and that to ceftazidime,cefepime and cefoperazone-sulbactam are relatively high.