Clinical characteristics and phylogenetic analysis of human enteric adenovirus type 41 (HAdV-F41) from children with gastroenteritis during SARS-CoV-2 pandemic.

Human adenovirus type 41 (HAdV-F41) usually causes pediatrics gastroenteritis. However, it was reported to be associated with the outbreaks of severe acute hepatitis of unknown aetiology (SAHUA) in pediatrics during COVID-19 pandemic. In this study, we investigated the prevalence of enteric HAdV-F41 in 37,920 paediatric gastroenteritis cases from 2017 to 2022 in Guangzhou, China. All children presented were tested negative for SARS-CoV-2 during the "zero-COVID" period. The main clinical symptom of the children was diarrhea (96.5%). No fatalities nor liver abnormal symptoms was found. In 2021, one year since the pandemic of COVID-19, the prevalence of HAdV-F41 abruptly increased from 3.71% to 8.64% (P < 0.001). All of HAdV-F41 circulating worldwide were classified into eight different subtypes (G1-G8) based on the phylogenetic clustering permutation of the four capsid genes of HAdV-F41. G3 was the predominant subtype (56.2%; 77/137). CRV5 isolates from SAHUA cases belong to this subtype, in which N312D and H335D mutations in the short fiber knob were identified in both Guangzhou and CRV5 isolates, presumably changing the virus tropism by directly interacting with the heparin sulfate (HS) receptor. Additionally, a novel recombinant G6 subtype, which is unique and only circulating in China was first identified in this study. This is the first study highlighting the prevalence of HAdV-F41 in paediatric cases of gastroenteritis during COVID-19 pandemic in China. The clinical and viral evolution finding of HAdV-F41 provide insight into the clinical characteristics of children with HAdV-F41 infections as well as the uncertain role of HAdV-F41 in the cause of SAHUA.

Mouse KL2 is a unique MTSE involved in chromosome-based spindle organization and regulated by multiple kinases during female meiosis.

Microtubule-severing enzymes (MTSEs) play important roles in mitosis and meiosis of the primitive organisms. However, no studies have assessed their roles in mammalian meiosis of females, whose abnormality accounts for over 80% of the cases of gamete-originated human reproductive disease. In the current study, we reported that katanin-like 2 (KL2) was the only MTSE concentrating at chromosomes. Furthermore, the knockdown of KL2 significantly reduced chromosome-based increase in the microtubule (MT) polymer, increased aberrant kinetochore-MT (K-MT) attachment, delayed meiosis, and severely affected normal fertility. Importantly, we demonstrated that the inhibition of aurora B, a key kinase for correcting aberrant K-MT attachment, eliminated KL2 from chromosomes completely. KL2 also interacted with phosphorylated eukaryotic elongation factor-2 kinase; they competed for chromosome binding. We also observed that the phosphorylated KL2 was localized at spindle poles, and that KL2 phosphorylation was regulated by extracellular signal-regulated kinase 1/2. In summary, our study reveals a novel function of MTSEs in mammalian female meiosis and demonstrates that multiple kinases coordinate to regulate the levels of KL2 at chromosomes.

Oocytes could rearrange immunoglobulin production to survive over adverse environmental stimuli.

Immunoglobulins are key humoral immune molecules produced and secreted by B lymphocytes at various stages of differentiation. No research has reported whether immunoglobulins are present in the non-proliferative female germ cells-oocytes-and whether they are functionally important for oocyte quality, self-protection, and survival. Herein, we found that IgG was present in the oocytes of immunodeficient mice; the IgG-VDJ regions were highly variable between different oocytes, and H3K27Ac bound and regulated the IgG promoter region. Next, IgG mRNA and protein levels increased in response to LPS, and this increment was mediated by CR2 on the oocyte membrane. Finally, we revealed three aspects of the functional relevance of oocyte IgG: first, oocytes could upregulate IgG to counteract the increased ROS level induced by CSF1; second, oocytes could upregulate IgG in response to injected virus ssRNA to maintain mitochondrial integrity; third, upon bacterial infection, oocytes could secrete IgG, subsequently encompassing the bacteria, thus increasing survival compared to somatic cells. This study reveals for the first time that the female germ cells, oocytes, can independently adjust intrinsic IgG production to survive in adverse environments.

Impact of intergenerational support and medical expenditures on depression: Evidence from rural older adults in China.

Objective: Globally, depression has become a major health issue among older adults, who experience poor physical health and high medical expenditures. In Asian countries, older adults are greatly dependent on their children. This study assessed the impact of different types of intergenerational support and medical expenditures on depression among older adults in rural China. Method: A three-phase balanced panel was constructed based on data from 1,838 rural older adults with comparable scores on the Center for Epidemiologic Studies Depression Scale (CES-D) from the China Family Panel Studies in 2012, 2016, and 2018. A fixed-effects model was used to analyze the impact of intergenerational support and medical expenditures on CES-D score and of intergenerational support on medical expenditures. The propensity score-matching model was used to test the regression results' robustness. Results: The findings were as follows. First, different types of intergenerational support had a heterogeneous impact on depression among rural older adults. Emotional support had a significantly negative impact on CES-D score, although too much care-based support had a positive impact on CES-D score. Low-level economic support had no significant effect on CES-D score. Second, medical expenditures impacted depression; among these, non-inpatient medical expenditure had a significant and positive impact on CES-D score. Third, CES-D scores among rural older adults were associated with chronic diseases and per capita family income. Fourth, care-based support was associated with reduced non-inpatient medical expenditures, and the sub-sample regression results indicated that the impact was significant for older adults with no chronic diseases and those younger than 75 years. Conclusion: Intergenerational emotional support and non-inpatient medical expenditures directly affected rural older adults' CES-D scores. The mediating role of medical expenditures between intergenerational support and CES-D score was not significant. Measures should be taken to encourage intergenerational emotional support and reduce the pressure on children's economic and care-based support. Further, the medical insurance reimbursement policy, as formal support, should be improved to alleviate depression among rural older adults when children's support is limited.

Fidgetin knockdown and knockout influences female reproduction distinctly in mice.

Microtubule-severing proteins (MTSPs), are a family of proteins which use adenosine triphosphate to sever microtubules. MTSPs have been shown to play an important role in multiple microtubule-involved cellular processes. One member of this family, fidgetin ( FIGN), is also involved in male fertility; however, no studies have explored its roles in female fertility. In this study, we found mouse fidgetin is rich within oocyte zona pellucida (ZP) and is the only MTSP member to do so. Fidgetin also appears to interact with all three ZP proteins. These findings prompted us to propose that fidgetin might prevent polyspermy. Results from in vitro maturation oocytes analysis showed that fidgetin knockdown did cause polyspermy. We then deleted all three fidgetin isoforms with CRISPR/Cas9 technologies; however, female mice remained healthy and with normal fertility. Of all mouse MTSPs, only the mRNA level of fidgetin-like 1 ( FIGNL1) significantly increased. Therefore, we assert that fidgetin-like 1 compensates fidgetin's roles in fidgetin knockout female mice.

Development of Receptor Binding Domain (RBD)-Conjugated Nanoparticle Vaccines with Broad Neutralization against SARS-CoV-2 Delta and Other Variants.

The SARS-CoV-2 Delta (B.1.617.2) strain is a variant of concern (VOC) that has become the dominant strain worldwide in 2021. Its transmission capacity is approximately twice that of the original strain, with a shorter incubation period and higher viral load during infection. Importantly, the breakthrough infections of the Delta variant have continued to emerge in the first-generation vaccine recipients. There is thus an urgent need to develop a novel vaccine with SARS-CoV-2 variants as the major target. Here, receptor binding domain (RBD)-conjugated nanoparticle vaccines targeting the Delta variant, as well as the early and Beta/Gamma strains, are developed. Under both a single-dose and a prime-boost strategy, these RBD-conjugated nanoparticle vaccines induce the abundant neutralizing antibodies (NAbs) and significantly protect hACE2 mice from infection by the authentic SARS-CoV-2 Delta strain, as well as the early and Beta strains. Furthermore, the elicitation of the robust production of broader cross-protective NAbs against almost all the notable SARS-CoV-2 variants including the Omicron variant in rhesus macaques by the third re-boost with trivalent vaccines is found. These results suggest that RBD-based monovalent or multivalent nanoparticle vaccines provide a promising second-generation vaccine strategy for SARS-CoV-2 variants.

A bivalent nanoparticle vaccine exhibits potent cross-protection against the variants of SARS-CoV-2.

Inoculation against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is ongoing worldwide. However, the emergence of SARS-CoV-2 variants could cause immune evasion. We developed a bivalent nanoparticle vaccine that displays the receptor binding domains (RBDs) of the D614G and B.1.351 strains. With a prime-boost or a single-dose strategy, this vaccine elicits a robust neutralizing antibody and full protection against infection with the authentic D614G or B.1.351 strain in human angiotensin-converting enzyme 2 transgene mice. Interestingly, 8 months after inoculation with the D614G-specific vaccine, a new boost with this bivalent vaccine potently elicits cross-neutralizing antibodies for SARS-CoV-2 variants in rhesus macaques. We suggest that the D614G/B.1.351 bivalent vaccine could be used as an initial single dose or a sequential enforcement dose to prevent infection with SARS-CoV-2 and its variants.

Rapid detection of KPC-producing Klebsiella pneumoniae in China based on MALDI-TOF MS.

Klebsiella pneumoniae carbapenemase-producing K. pneumoniae (KPC-Kp) represent a serious threat to public health and their timely detection is essential for patient management and the prevention of nosocomial infections. Here, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used to rapidly identify dominant KPC-Kp in China, by using an automated detection of a KPC-specific peak (at 4521 m/z) by a genetic algorithm using ClinProTools software. Whole-genome sequencing (WGS) was used to understand the genetic environment of the blaKPC-2 gene. In this study, we analyzed 235 K. pneumoniae Chinese clinical isolates, of which 175 (93 KPC-positive isolates and 82 KPC-negative isolates) isolates were used to build a model to select a KPC-specific peak, and another 60 isolates for external validation. In addition, all the spectra were visually inspected by the FlexAnalysis software to evaluate the accuracy of the automated detection. The results showed a 4521 m/z peak found in all blaKPC-2-positive isolates but absent in blaKPC-2-negative isolates. Interestingly, all KPC-Kp belonged to ST11, the dominant clone in China. WGS analysis of a representative isolate showed that the genetic environment of KPC-2 was IS26-ISKpn27-blaKPC-2-ΔISKpn6-Tn1721, similar to the KPC-2 genetic environment of ST11 KPC-Kp previously reported in China. Therefore, the 4521 m/z peak is closely related to ST11 KPC-Kp. In summary, we used MALDI-TOF MS to quickly detect KPC-Kp in the process of routine bacterial identification without increasing costs or requiring further knowledge, which has broad application prospects in drug resistance analysis and infection control.

Gm364 coordinates MIB2/DLL3/Notch2 to regulate female fertility through AKT activation.

Many integral membrane proteins might act as indispensable coordinators in specific functional microdomains to maintain the normal operation of known receptors, such as Notch. Gm364 is a multi-pass transmembrane protein that has been screened as a potential female fertility factor. However, there have been no reports to date about its function in female fertility. Here, we found that global knockout of Gm364 decreased the numbers of primordial follicles and growing follicles, impaired oocyte quality as indicated by increased ROS and γ-H2AX, decreased mitochondrial membrane potential, decreased oocyte maturation, and increased aneuploidy. Mechanistically, Gm364 directly binds and anchors MIB2, a ubiquitin ligase, on the membrane. Subsequently, membrane MIB2 ubiquitinates and activates DLL3. Next, the activated DLL3 binds and activates Notch2, which is subsequently cleaved within the cytoplasm to produce NICD2, the intracellular active domain of Notch2. Finally, NICD2 can directly activate AKT within the cytoplasm to regulate oocyte meiosis and quality.

ATP5O Hypo-crotonylation Caused by HDAC2 Hyper-Phosphorylation Is a Primary Detrimental Factor for Downregulated Phospholipid Metabolism under Chronic Stress.

Objective. Chronic stress (CS)-induced abnormal metabolism and other subsequent aspects of abnormality are threatening human health. Little is known regarding whether and how protein post-translational-modifications (PTMs) correlate with abnormal metabolism under CS. The aim of this study was to address this issue and also identify novel key protein PTM. Methods. First, we screened which pan-PTM had significant change between control and CS female mice and whether clinical CS females had similar pan-PTM change. Second, we performed quantitative PTM-omics and metabolomics to verify the correlation between abnormal protein PTMs and atypical metabolism. Third, we performed quantitative phospho-omics to identify the key PTM-regulating enzyme and investigate the interaction between PTM protein and PTM-regulating enzyme. Fourth, we attempted to rectify the abnormal metabolism by correcting the activity of the PTM-regulating enzyme. Finally, we examined whether the selected key protein was also correlated with stress scores and atypical metabolism in clinical women. Results. We initially found that multiple tissues of CS female mice have downregulated pan-crotonylation, and verified that the plasma of clinical CS females also had downregulated pan-crotonylation. Then we determined that ATP5O-K51 crotonylation decreased the most and also caused gross ATP5O decrement, whereas the plasma of CS mice had downregulated phospholipids. Next, downregulating ATP5O crotonylation partially recapitulated the downregulated phospholipid metabolism in CS mice. Next, we verified that HDAC2-S424 phosphorylation determined its decrotonylation activity on ATP5O-K51. Furthermore, correcting HDAC2 hyper-phosphorylation recovered the gross ATP5O level and partially rescued the downregulated phospholipid metabolism in CS mice. Finally, the ATP5O level was also significantly lower and correlated with high stress scores and downregulated phospholipid metabolism in clinical female plasma. Conclusion. This study discovered a novel PTM mechanism involving two distinct types of PTM in CS and provided a novel reference for the clinical precautions and treatments of CS.

GTPases Arf5 and Arl2 function partially distinctly during oocyte meiosis.

Mammalian female meiosis must be tightly regulated to produce high-quality mature oocytes for subsequent regular fertilization and healthy live birth of the next generation. GTPases control many important signal pathways involved in diverse cellular activities. ADP-ribosylation factor family members (Arfs) in mice possess GTPase activities, and some members have been found to function in meiosis. However, whether other Arfs play a role in meiosis is unknown. In this study, we found that Arl2 and Arf5 are the richest among Arfs in mouse oocytes, and they are more abundant in oocytes than in granular cells. Furthermore, Arl2 and Arf5 depletion both impeded meiotic progression, but by affecting spindles and microfilaments, respectively. Moreover, Arl2 and Arf5 depletion both significantly increased regular reactive oxygen species levels and decreased mitochondrial membrane potential and autophagy, indicating that oocyte quality was damaged by Arl2 and Arf5 depletion. These results suggest that Arl2 and Arf5 are two novel essential GTPases required for oocyte meiosis and quality control.

Rat BAT xenotransplantation recovers the fertility and metabolic health of PCOS mice.

Polycystic ovarian syndrome (PCOS) is a major severe ovary disorder affecting 5-10% of reproductive women around the world. PCOS can be considered a metabolic disease because it is often accompanied by obesity and diabetes. Brown adipose tissue (BAT) contains abundant mitochondria and adipokines and has been proven to be effective for treating various metabolic diseases. Recently, allotransplanted BAT successfully recovered the ovarian function of PCOS rat. However, BAT allotransplantation could not be applied to human PCOS; the most potent BAT is from infants, so voluntary donors are almost inaccessible. We recently reported that single BAT xenotransplantation significantly prolonged the fertility of aging mice and did not cause obvious immunorejection. However, PCOS individuals have distinct physiologies from aging mice; thus, it remains essential to study whether xenotransplanted rat BAT can be used for treating PCOS mice. In this study, rat-to-mouse BAT xenotransplantation, fortunately, did not cause severe rejection reaction, and significantly recovered ovarian functions, indicated by the recovery of fertility, oocyte quality, and the levels of multiple essential genes and kinases. Besides, the blood biochemical index, glucose resistance, and insulin resistance were improved. Moreover, transcriptome analysis showed that the recovered PCOS F0 mother following BAT xenotransplantation could also benefit the F1 generation. Finally, BAT xenotransplantation corrected characteristic gene expression abnormalities found in the ovaries of human PCOS patients. These findings suggest that BAT xenotransplantation could be a novel therapeutic strategy for treating PCOS patients.

MALDI-TOF MS platform combined with machine learning to establish a model for rapid identification of methicillin-resistant Staphylococcus aureus.

MALDI-TOF MS is an effective potential tool to distinguish between MSSA and MRSA. By combining the ClinProTools3.0 software and manual grouping intervention, we proposed a model optimization method for the first time. The cross validation of the model increased from 95.82% to 96.68%, and the accuracy of the model increased from 88.89% to 91.98%. Finally, we reported nine characteristic peaks of rapid detection of MRSA.

Multilocus Sequence Typing Reveals Clonality of Fluconazole-Nonsusceptible Candida tropicalis: A Study From Wuhan to the Global.

Candida tropicalis is a globally distributed human pathogenic yeast, and its increasing resistance to azoles makes clinical treatment difficult. In this study, we investigated the clinical features, azole resistance and genetic relatedness of 87 C. tropicalis isolates from central China and combined with the global database to explore the relationship between genetic information and fluconazole susceptibility. Of the 55 diploid sequence types (DSTs) identified by multilocus sequence typing (MLST), 27 DSTs were new to the C. tropicalis MLST database. Fluconazole-nonsusceptible (FNS) isolates were genetically closely related. goeBURST analysis showed that DST225, DST376, DST506, and DST546 formed a distinct and unique FNS clonal complex (CC) in Wuhan. The local FNS CC belongs to the large FNS CC (CC2) in China, in which the putative founder DST225 has been reported from the environment. The three most prevalent types (DST506, DST525, and DST546) in Wuhan had high minimum inhibitory concentrations (MICs) for antifungal azoles, and the six possible nosocomial transmissions we captured were all FNS strains, most of which were from CC2. Unique FNS CCs have been found in Singapore (CC8) and India (CC17) and are close to China's CC2 in the minimum spanning tree. There were no FNS CCs outside Asia. This study is the first to reveal a significant correlation between genetic information and fluconazole susceptibility worldwide and to trace geographical locations, which is of great value for molecular epidemiological surveillance and azole-resistance study of C. tropicalis globally.

Downregulation of the ubiquitin ligase KBTBD8 prevented epithelial ovarian cancer progression.

OBJECTIVES: Kelch repeat and BTB domain-containing protein 8, KBTBD8, has been identified as a female fertility factor. However, there have been no reports on the role of KBTBD8 in the progression of epithelial ovarian cancer, EOC. Our study aimed to address this issue. METHODS: We first examine KBTBD8 expression in EOC tissues and cells. Next, we performed RNA sequencing to reveal the overall mechanism. Then we investigated the roles of KBTBD8 in the proliferation, migration, and health status of cultured EOC cells. Finally, we employed tumor xenograft models to evaluate the role of KBTBD8 in vivo. RESULTS: First, KBTBD8 level was significantly higher in EOC tissues and cells. Next, comparative RNA sequencing identified more tumorigenesis-related genes that KBTBD8 might regulate. Then we found that KBTBD8 knockdown significantly decreased EOC cell proliferation, migration, and the activities of multiple tumorigenesis-related kinases. Finally, KBTBD8 knockdown significantly diminished ovarian tumor formation in vivo. CONCLUSION: Proper KBTBD8 level is essential for the healthy growth of ovarian somatic cells, such as ovarian epithelial cells. Excessive KBTBD8 might be a significant impetus for EOC progression. KBTBD8 reduction greatly inhibits EOC proliferation and migration.

Molecular epidemiology of Candida tropicalis isolated from urogenital tract infections.

Candida tropicalis is a common human pathogenic yeast, and its molecular typing is important for studying the population structure and epidemiology of this opportunistic yeast, such as epidemic genotype, population dynamics, nosocomial infection, and drug resistance surveillance. In this study, the antifungal susceptibility test and multilocus sequence typing (MLST) analysis were carried out on C. tropicalis from central China. Among 64 urogenital isolates, 45 diploid sequence types (DST) were found, of which 20 DSTs (44.4%) were new to the central database. The goeBURST analysis showed that CC1 (clonal complex) was the only azole-resistant (100%, 10/10) cluster in Wuhan, which was composed of DST546, DST225, DST376, and DST506, and most of the strains (90%, 9/10) were isolated from the urinary tract. Potential nosocomial infections were mainly caused by CC1 strains. The azole resistance rate of urinary isolates (50.0%, 21/42) was higher than that of vaginal isolates (27.3%, 6/22). The genotype diversity and novelty of vaginal isolates were higher than those of urinary isolates. C. tropicalis population in Wuhan was genetically diverse and divergent from that seen in other countries. In this study, there were significant differences in genotype and azole susceptibility between urine and vaginal strains. The azole-resistant cluster (CC1) found in urine is of great significance for the clinical treatment and prevention of nosocomial infection. The newly discovered DSTs will contribute to further study the similarity, genetic relationship, and molecular epidemiology of C. tropicalis worldwide.

Fam70A binds Wnt5a to regulate meiosis and quality of mouse oocytes.

OBJECTIVES: Little is known about the roles of integral membrane proteins beyond channels, carriers or receptors in meiotic oocytes. The transmembrane protein Fam70A was previously identified as a likely "female fertility factor" in Fox3a-knockout mouse ovaries where almost all follicles underwent synchronous activation and the mice became infertile very early. However, whether Fam70A functions in oocyte meiosis remains unknown. Therefore, the present study aimed to address this question. MATERIALS AND METHODS: Co-immunoprecipitation, immunogold labelling-electron microscopy, co-localization and yeast two-hybrid assays were used to verify the interaction. Antibody or small interfering RNA transfection was used to deplete the proteins. Immunofluorescence, immunohistochemistry and live tracker staining were used to examine the localization or characterize phenotypes. Western blot was used to examine the protein level. RESULTS: Fam70A was enriched in oocyte membranes important for normal meiosis. Fam70A depletion remarkably disrupted spindle assembly, chromosome congression and first polar body extrusion, which subsequently increased aneuploidy and abnormal fertilization. Moreover, Fam70A directly bound Wnt5a, the most abundant Wnt member within oocytes. Depletion of either Fam70A or Wnt5a remarkably increased adenomatous polyposis coli (APC), which stabilizes active ß-catenin and microtubules. Consequently, depletion of either Fam70A or Wnt5a remarkably increased p-ß-catenin (inactive form) and acetylated tubulin, while APC knockdown remarkably decreased these two. Furthermore, Fam70A depletion remarkably reduced Akt phosphorylation. CONCLUSIONS: Fam70A regulates meiosis and quality of mouse oocytes through both canonical and non-canonical Wnt5a signalling pathways.

[Bacteriologic profile and drug resistance in children with respiratory infection from 2016 to 2018].

OBJECTIVE: To study the bacteriologic profile and drug resistance of respiratory infection in children, and to provide a basis for etiological diagnosis and rational use of antimicrobial agents. METHODS: A retrospective analysis was performed for 15 047 children who attended the hospital due to respiratory infection from January 2016 to December 2018. Their sputum samples were collected, and the Phoenix-100 automatic microbial identification system was used for the identification and drug sensitivity analysis of the isolated pathogenic bacteria. RESULTS: Of all 17 174 sputum samples detected, there were 2 395 positive samples, with a positive rate of 13.95%; a total of 2 584 strains of pathogenic bacteria were isolated, among which there were 1 577 (61.03%) Gram-negative strains, 967 (37.42%) Gram-positive strains, and 40 (1.55%) fungal strains. The most common pathogen was Haemophilus influenzae (33.90%), followed by Streptococcus pneumoniae (33.55%), Moraxella catarrhalis (19.20%), and Staphylococcus aureus (3.64%). Among the 2 331 children with positive infection, 251 had mixed infection, most commonly with Haemophilus influenzae and Streptococcus pneumoniae. The detection rate of pathogenic bacteria was higher in winter and spring and lower in summer and autumn. There was a significant difference in the detection rate of pathogenic bacteria between different age groups (P<0.05), with the highest detection rate in infants aged 1 month to <1 year. Streptococcus pneumoniae and Staphylococcus aureus had a sensitivity rate of 100% to vancomycin, linezolid, and teicoplanin, and Haemophilus influenzae had a lower sensitivity rate to ampicillin, compound sulfamethoxazole and cefuroxime and a higher sensitivity rate to other drugs. CONCLUSIONS: Haemophilus influenzae, Streptococcus pneumoniae, and Moraxella catarrhalis are the main pathogenic bacteria of respiratory infection in children, and mixed infection is the most common type of infection. The detection rate of pathogenic bacteria varies across seasons and ages. Different pathogenic bacteria have different features of drug resistance, and antibiotics should be selected based on drug sensitivity results.

Performance of modified carbapenem inactivation method and inhibitor-based combined disk test in the detection and distinguishing of carbapenemase producing Enterobacteriaceae.

BACKGROUND: This study aimed to evaluate the performance of modified carbapenem inactivation method (mCIM) combined EDTA-carbapenem inactivation method (eCIM), and inhibitor-based combined disk test (CDT) in the detection and distinguishing of carbapenemase production in Enterobacteriaceae. METHODS: A total of 101 nonrepetitive carbapenem insensitive Enterobacteriaceae [minimal inhibitory concentration (MIC) ≥2 µg/mL] were tested by mCIM, eCIM and CDT respectively, and the major carbapenemase genes including blaKPC, blaNDM, blaIMP, blaVIM and blaOXA-48-like genes were detected by polymerase chain reaction (PCR) as control. RESULTS: Seventy-nine (78.2%) of isolates were found to harbour one or more carbapenemase genes by PCR, with blaKPC and blaNDM being the most common genes. OXA-48-like genes were undetectable. The coincidence rate of mCIM combined eCIM and CDT was 97.5% (77/79) and 96.2% (76/79) respectively, compared with gene detection. Both assays had a misclassification in two blaKPC+NDM-producing isolates of Klebsiella oxytoca. The sensitivity and specificity of two assays above were 100.0% vs. 95.0% and 98.4% vs. 98.4%, respectively in distinguishing serine-carbapenemase, while they were 95.1% vs. 97.6% and 100% vs. 100.0%, respectively in distinguishing metallo-carbapenemase. CONCLUSIONS: mCIM combined eCIM and the CDT are both useful tools for the reliable detection and distinguishing single serine-carbapenemase or metallo-carbapenemase, but not for mixed types.