Recently,inductively coupled plasma mass spectrometry(ICP-MS)has shown great potential in the quantitative analysis of biomolecules due to its high sensitivity,high accuracy,anti-interference ability and multi-element detection capability.Elemental tags enable specific labeling of target biomolecules with element-reporter groups,playing a critical role in the performance of bioanalysis using ICP-MS.Antibodies are highly specific to antigens on cell surfaces and are one of the most commonly used targeted delivery vehicles.Traditional random chemical coupling methods are difficult to control the coupling sites and loading numbers of element reporters,which lead to heterogeneity of antibody conjugates,affecting the specific recognition and binding ability of antibodies and antigens,and thus reducing the targeting ability and quantitative accuracy.To solve these issues,site-specific coupling of antibodies with element reporter was studied in this work.By using peptide-mediated affinity labeling technology,a short chain DNA was specifically labeled to immunoglobulin G(IgG)antibody K248,and then DNA base complementary pairing was used to realize accurate assembly of monoclonal antibodies and Eu-modified phage MS2 capsid,resulting in the preparation of highly sensitive and selective element tags.This method overcame the heterogeneity problem of element tags caused by random coupling,and ensured the targeting specificity,affinity and signal amplification ability of element tags.The element tag prepared here showed a specificity to bind/label with epidermal growth factor receptor(EGFR)biomarker on the surface of cancer cells,realizing single cancer cell analysis using ICP-MS.

Objective To investigate the clinical characteristics and prognosis of pneumococcal meningitis(PM),and drug sensitivity of Streptococcus pneumoniae(SP)isolates in Chinese children.Methods A retrospective analysis was conducted on clinical information,laboratory data,and microbiological data of 160 hospitalized children under 15 years old with PM from January 2019 to December 2020 in 33 tertiary hospitals across the country.Results Among the 160 children with PM,there were 103 males and 57 females.The age ranged from 15 days to 15 years,with 109 cases(68.1% )aged 3 months to under 3 years.SP strains were isolated from 95 cases(59.4% )in cerebrospinal fluid cultures and from 57 cases(35.6% )in blood cultures.The positive rates of SP detection by cerebrospinal fluid metagenomic next-generation sequencing and cerebrospinal fluid SP antigen testing were 40% (35/87)and 27% (21/78),respectively.Fifty-five cases(34.4% )had one or more risk factors for purulent meningitis,113 cases(70.6% )had one or more extra-cranial infectious foci,and 18 cases(11.3% )had underlying diseases.The most common clinical symptoms were fever(147 cases,91.9% ),followed by lethargy(98 cases,61.3% )and vomiting(61 cases,38.1% ).Sixty-nine cases(43.1% )experienced intracranial complications during hospitalization,with subdural effusion and/or empyema being the most common complication[43 cases(26.9% )],followed by hydrocephalus in 24 cases(15.0% ),brain abscess in 23 cases(14.4% ),and cerebral hemorrhage in 8 cases(5.0% ).Subdural effusion and/or empyema and hydrocephalus mainly occurred in children under 1 year old,with rates of 91% (39/43)and 83% (20/24),respectively.SP strains exhibited complete sensitivity to vancomycin(100% ,75/75),linezolid(100% ,56/56),and meropenem(100% ,6/6).High sensitivity rates were also observed for levofloxacin(81% ,22/27),moxifloxacin(82% ,14/17),rifampicin(96% ,25/26),and chloramphenicol(91% ,21/23).However,low sensitivity rates were found for penicillin(16% ,11/68)and clindamycin(6% ,1/17),and SP strains were completely resistant to erythromycin(100% ,31/31).The rates of discharge with cure and improvement were 22.5% (36/160)and 66.2% (106/160),respectively,while 18 cases(11.3% )had adverse outcomes.Conclusions Pediatric PM is more common in children aged 3 months to under 3 years.Intracranial complications are more frequently observed in children under 1 year old.Fever is the most common clinical manifestation of PM,and subdural effusion/emphysema and hydrocephalus are the most frequent complications.Non-culture detection methods for cerebrospinal fluid can improve pathogen detection rates.Adverse outcomes can be noted in more than 10% of PM cases.SP strains are high sensitivity to vancomycin,linezolid,meropenem,levofloxacin,moxifloxacin,rifampicin,and chloramphenicol.[Chinese Journal of Contemporary Pediatrics,2024,26(2):131-138]

A quartz crystal microbalance(QCM)-systematic evolution of ligands by the exponential en-richment(SELEX)technique was developed to screen out aptamers with high affinity for arsenic(Ⅲ).A random single strand DNA library was designed and fixed on the mercaptoethylamine-modified crystal plate with arsenic(Ⅲ)as the target,and the free aptamer was captured in the solution,and the QCM-SELEX screening method was constructed.After 6 rounds of screening,the secondary library was se-quenced with high throughput method,and the 6S1 dissociation coefficient Kd value was 0.36 μmol/L based on QCM resonance frequency.Using 6S1 as a probe,the QCM biosensor was constructed for the detection of arsenic(Ⅲ).The sensor has a good linear relationship in the range of 0.01 μmol/L～0.2μmol/L,and the detection limit of arsenic(Ⅲ)is 5.2 nmol/L(3σ),indicatinggood selectivity.

Acanthamoeba is an opportunistic pathogen that causes Acanthamoeba keratitis, granulomatous amoebic encephalitis, and other cutaneous diseases. The life cycle of Acanthamoeba consists of 2 stages of trophozoites and cysts. Under adverse environmental conditions, Acanthamoeba encysts, while the conditions become favorable for growth, it reverts to the trophozoite form. Acanthamoeba excystation is crucial for its proliferation and can lead to recurrent infections after incomplete treatment. To identify the factors involved in excystation, A. castellanii was subjected to either encystation- or excystation-inducing conditions, and gene expression profiles were compared using mRNA sequencing. A. castellanii samples were collected at 8 h intervals for analysis under both conditions. Differentially expressed gene analysis revealed that 1,214 and 1,163 genes were upregulated and downregulated, respectively, by more than 2-fold during early excystation. Five genes markedly upregulated in early excystation (ACA1_031140, ACA1_032330, ACA1_374400, ACA1_275740, and ACA1_112650) were selected, and their expression levels were confirmed via real-time PCR. Small interfering RNA (siRNA) targeting these 5 genes was transfected into Acanthamoeba and gene knockdown was validated through real-time PCR. The silencing of ACA1_031140, ACA1_032330, ACA1_374400, and ACA1_112650 inhibited excystation and suggested that these genes might be essential for excystation. Our findings provide valuable insights for suppressing Acanthamoeba proliferation and recurrence.

Acanthamoeba is an opportunistic pathogen that causes Acanthamoeba keratitis, granulomatous amoebic encephalitis, and other cutaneous diseases. The life cycle of Acanthamoeba consists of 2 stages of trophozoites and cysts. Under adverse environmental conditions, Acanthamoeba encysts, while the conditions become favorable for growth, it reverts to the trophozoite form. Acanthamoeba excystation is crucial for its proliferation and can lead to recurrent infections after incomplete treatment. To identify the factors involved in excystation, A. castellanii was subjected to either encystation- or excystation-inducing conditions, and gene expression profiles were compared using mRNA sequencing. A. castellanii samples were collected at 8 h intervals for analysis under both conditions. Differentially expressed gene analysis revealed that 1,214 and 1,163 genes were upregulated and downregulated, respectively, by more than 2-fold during early excystation. Five genes markedly upregulated in early excystation (ACA1_031140, ACA1_032330, ACA1_374400, ACA1_275740, and ACA1_112650) were selected, and their expression levels were confirmed via real-time PCR. Small interfering RNA (siRNA) targeting these 5 genes was transfected into Acanthamoeba and gene knockdown was validated through real-time PCR. The silencing of ACA1_031140, ACA1_032330, ACA1_374400, and ACA1_112650 inhibited excystation and suggested that these genes might be essential for excystation. Our findings provide valuable insights for suppressing Acanthamoeba proliferation and recurrence.

Acanthamoeba is an opportunistic pathogen that causes Acanthamoeba keratitis, granulomatous amoebic encephalitis, and other cutaneous diseases. The life cycle of Acanthamoeba consists of 2 stages of trophozoites and cysts. Under adverse environmental conditions, Acanthamoeba encysts, while the conditions become favorable for growth, it reverts to the trophozoite form. Acanthamoeba excystation is crucial for its proliferation and can lead to recurrent infections after incomplete treatment. To identify the factors involved in excystation, A. castellanii was subjected to either encystation- or excystation-inducing conditions, and gene expression profiles were compared using mRNA sequencing. A. castellanii samples were collected at 8 h intervals for analysis under both conditions. Differentially expressed gene analysis revealed that 1,214 and 1,163 genes were upregulated and downregulated, respectively, by more than 2-fold during early excystation. Five genes markedly upregulated in early excystation (ACA1_031140, ACA1_032330, ACA1_374400, ACA1_275740, and ACA1_112650) were selected, and their expression levels were confirmed via real-time PCR. Small interfering RNA (siRNA) targeting these 5 genes was transfected into Acanthamoeba and gene knockdown was validated through real-time PCR. The silencing of ACA1_031140, ACA1_032330, ACA1_374400, and ACA1_112650 inhibited excystation and suggested that these genes might be essential for excystation. Our findings provide valuable insights for suppressing Acanthamoeba proliferation and recurrence.

Acanthamoeba is an opportunistic pathogen that causes Acanthamoeba keratitis, granulomatous amoebic encephalitis, and other cutaneous diseases. The life cycle of Acanthamoeba consists of 2 stages of trophozoites and cysts. Under adverse environmental conditions, Acanthamoeba encysts, while the conditions become favorable for growth, it reverts to the trophozoite form. Acanthamoeba excystation is crucial for its proliferation and can lead to recurrent infections after incomplete treatment. To identify the factors involved in excystation, A. castellanii was subjected to either encystation- or excystation-inducing conditions, and gene expression profiles were compared using mRNA sequencing. A. castellanii samples were collected at 8 h intervals for analysis under both conditions. Differentially expressed gene analysis revealed that 1,214 and 1,163 genes were upregulated and downregulated, respectively, by more than 2-fold during early excystation. Five genes markedly upregulated in early excystation (ACA1_031140, ACA1_032330, ACA1_374400, ACA1_275740, and ACA1_112650) were selected, and their expression levels were confirmed via real-time PCR. Small interfering RNA (siRNA) targeting these 5 genes was transfected into Acanthamoeba and gene knockdown was validated through real-time PCR. The silencing of ACA1_031140, ACA1_032330, ACA1_374400, and ACA1_112650 inhibited excystation and suggested that these genes might be essential for excystation. Our findings provide valuable insights for suppressing Acanthamoeba proliferation and recurrence.

Acanthamoeba is an opportunistic pathogen that causes Acanthamoeba keratitis, granulomatous amoebic encephalitis, and other cutaneous diseases. The life cycle of Acanthamoeba consists of 2 stages of trophozoites and cysts. Under adverse environmental conditions, Acanthamoeba encysts, while the conditions become favorable for growth, it reverts to the trophozoite form. Acanthamoeba excystation is crucial for its proliferation and can lead to recurrent infections after incomplete treatment. To identify the factors involved in excystation, A. castellanii was subjected to either encystation- or excystation-inducing conditions, and gene expression profiles were compared using mRNA sequencing. A. castellanii samples were collected at 8 h intervals for analysis under both conditions. Differentially expressed gene analysis revealed that 1,214 and 1,163 genes were upregulated and downregulated, respectively, by more than 2-fold during early excystation. Five genes markedly upregulated in early excystation (ACA1_031140, ACA1_032330, ACA1_374400, ACA1_275740, and ACA1_112650) were selected, and their expression levels were confirmed via real-time PCR. Small interfering RNA (siRNA) targeting these 5 genes was transfected into Acanthamoeba and gene knockdown was validated through real-time PCR. The silencing of ACA1_031140, ACA1_032330, ACA1_374400, and ACA1_112650 inhibited excystation and suggested that these genes might be essential for excystation. Our findings provide valuable insights for suppressing Acanthamoeba proliferation and recurrence.

Malaria is a global disease affecting a large portion of the world’s population. Although vaccines have recently become available, their efficacies are suboptimal. We generated virus-like particles (VLPs) that expressed either apical membrane antigen 1 (AMA1) or microneme-associated antigen (MIC) of Plasmodium berghei and compared their efficacy in BALB/c mice. We found that immune sera acquired from AMA1 VLP- or MIC VLP-immunized mice specifically interacted with the antigen of choice and the whole P. berghei lysate antigen, indicating that the antibodies were highly parasite-specific. Both VLP vaccines significantly enhanced germinal center B cell frequencies in the inguinal lymph nodes of mice compared with the control, but only the mice that received MIC VLPs showed significantly enhanced CD4+ T cell responses in the blood following P. berghei challenge infection. AMA1 and MIC VLPs significantly suppressed TNF-α and interleukin-10 production but had a negligible effect on interferon-γ. Both VLPs prevented excessive parasitemia buildup in immunized mice, although parasite burden reduction induced by MIC VLPs was slightly more effective than that induced by AMA1. Both VLPs were equally effective at preventing body weight loss. Our findings demonstrated that the MIC VLP was an effective inducer of protection against murine experimental malaria and should be the focus of further development.

Objective:To investigate the diagnostic accuracy of serological indicators and evaluate the diagnostic value of a new established combined serological model on identifying the minimal hepatic encephalopathy (MHE) in patients with compensated cirrhosis.Methods:This prospective multicenter study enrolled 263 compensated cirrhotic patients from 23 hospitals in 15 provinces, autonomous regions and municipalities of China between October 2021 and August 2022. Clinical data and laboratory test results were collected, and the model for end-stage liver disease (MELD) score was calculated. Ammonia level was corrected to the upper limit of normal (AMM-ULN) by the baseline blood ammonia measurements/upper limit of the normal reference value. MHE was diagnosed by combined abnormal number connection test-A and abnormal digit symbol test as suggested by Guidelines on the management of hepatic encephalopathy in cirrhosis. The patients were randomly divided (7∶3) into training set ( n=185) and validation set ( n=78) based on caret package of R language. Logistic regression was used to establish a combined model of MHE diagnosis. The diagnostic performance was evaluated by the area under the curve (AUC) of receiver operating characteristic curve, Hosmer-Lemeshow test and calibration curve. The internal verification was carried out by the Bootstrap method ( n=200). AUC comparisons were achieved using the Delong test. Results:In the training set, prevalence of MHE was 37.8% (70/185). There were statistically significant differences in AMM-ULN, albumin, platelet, alkaline phosphatase, international normalized ratio, MELD score and education between non-MHE group and MHE group (all P<0.05). Multivariate Logistic regression analysis showed that AMM-ULN [odds ratio ( OR)=1.78, 95% confidence interval ( CI) 1.05-3.14, P=0.038] and MELD score ( OR=1.11, 95% CI 1.04-1.20, P=0.002) were independent risk factors for MHE, and the AUC for predicting MHE were 0.663, 0.625, respectively. Compared with the use of blood AMM-ULN and MELD score alone, the AUC of the combined model of AMM-ULN, MELD score and education exhibited better predictive performance in determining the presence of MHE was 0.755, the specificity and sensitivity was 85.2% and 55.7%, respectively. Hosmer-Lemeshow test and calibration curve showed that the model had good calibration ( P=0.733). The AUC for internal validation of the combined model for diagnosing MHE was 0.752. In the validation set, the AUC of the combined model for diagnosing MHE was 0.794, and Hosmer-Lemeshow test showed good calibration ( P=0.841). Conclusion:Use of the combined model including AMM-ULN, MELD score and education could improve the predictive efficiency of MHE among patients with compensated cirrhosis.