Objective To investigate the temporal changes in pathological damage and macrophage polarization in liver and spleen tissues of mice infected with Angiostrongylus cantonensis, and to preliminarily unravel the peripheral immune responses during the early stage of A. cantonensis infection. Methods Forty female BALB/c mice at ages of 6 to 8 weeks were randomly divided into four groups, including the control group and 7-, 14-, and 21-day infection groups, with 10 mice in each group. Each mouse in the infection groups was inoculated with 30 third-stage (L3) larvae of A. cantonensis by oral gavage, and five mice were randomly selected from each infection group on days 7, 14, and 21 post-infection, while mice in the control group were given the same volume of physiological saline and five mice were randomly selected from the control group on the day of oral gavage. Mouse liver and spleen tissues were sampled. The histopathological changes of mouse liver and spleen tissues were observed using hematoxylin and eosin (HE) staining, and the percentage of positive staining area and the co-localization positive rates of the macrophage surface antigens F4/80, CD86, and CD206 were quantified in mouse liver and spleen tissues using immunohistochemical and immunofluorescence staining. In addition, five mice were collected from each infection group on days 7, 14, and 21 post-infection, and five mice were collected from the control group on the day of oral gavage. Mouse liver and spleen tissues were sampled for detection of macrophage markers CD86 and CD206 and macrophage phenotyping using flow cytometry, and the expression of M1 macrophage markers, including inducible nitric oxide synthase (Nos2), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and M2 markers, including arginase 1 (Arg1), mannose receptor C-type 1 (Mrc1) and chitinase-like protein 3 (Chil3) was quantified in mouse liver and spleen tissues using real-time quantitative PCR (RT-qPCR) assay. Results Proliferative lesions of the hepatocyte were observed in mouse liver tissues and the follicular structures of the mouse spleen white pulp were disrupted 21 days post-infection with A. cantonensis. Immunohistochemical staining showed that there were significant differences in the percentages of F4/80, CD86 and CD206 positive staining areas in the liver and spleen tissues among the four groups of mice (F = 242.40, 197.14, 183.19, 157.65, 242.35 and 146.24; all P values < 0.001), and the percentages of positive staining in the liver and spleen tissues of mice in the 14-day infection group [(4.45 ± 0.51)%, (3.74 ± 0.67)%, (8.32 ± 0.72)%, (16.56 ± 1.14)%, (11.62 ± 0.52)%, and (8.29 ± 0.72)%, respectively] and the 21-day infection group [(3.70 ± 0.11)%, (3.22 ± 0.43)%, (11.53 ± 1.03)%, (12.59 ± 1.05)%, (9.02 ± 0.83)%, and (11.67 ± 1.10)%, respectively] were higher than in the control group [(0.35 ± 0.16)%, (0.40 ± 0.02)%, (0.93 ± 0.05)%, (2.78 ± 0.26)%, (2.33 ± 0.20)%, and (1.85 ± 0.20)%, respectively] (all P values < 0.05). Immunofluorescence staining showed significant differences in the positive rates of F4/80 co-localization with CD86 and CD206 in mouse liver and spleen tissues among the four groups (F = 24.42, 25.28, 54.51 and 130.55; all P values < 0.001). Flow cytometry detected significant differences in the proportions of CD86⁺ and CD206⁺ macrophages in mouse liver and spleen tissues among the four groups (F = 67.98, 18.41, 29.77, 172.80; all P values < 0.001), and the proportions of CD206⁺ macrophages in the liver and spleen of the 21-day infection group were significantly higher than those in the control group [(9.25 ± 2.55)% vs (3.83 ± 0.72)%, and (4.22 ± 0.56)% vs (0.47 ± 0.18)%, respectively] (both P values < 0.05). In addition, RT-qPCR assay quantified significant differences in the relative mRNA expression of M1 macrophage markers (IL-1β, TNF-α and Nos2) and M2 macrophage markers (Arg1, Chil3 and Mrc1) in mouse liver and spleen tissues among the four groups (F = 41.30, 31.82, 199.33, 19.96, 62.01, 119.76, 23.67, 95.90, 72.27, 82.59, 123.41 and 29.75; all P values < 0.05). Conclusions A. cantonensis infection may cause progressive pathological damage in mouse liver and spleen tissues, accompanied by dynamic temporal changes in macrophage polarization. M1 macrophage polarization predominates at the early stage of A. cantonensis infection and shifts towards M2 polarization at the later stages, suggesting that M2 polarization may participate in immune regulation at late stages of A. cantonensis infection by suppressing excessive inflammatory responses and promoting tissue repair.

Objective To investigate the changes in eosinophil counts and the activation of microglial cells in the brain tissues of mice at different stages of Angiostrongylus cantonensis infection, and to examine the role of microglia in regulating the progression of angiostrongyliasis and unravel the possible molecular mechanisms. Methods Fifty BALB/c mice were randomly divided into the control group and the 7-d, 14-d, 21-day and 25-d infection groups, of 10 mice in each group. All mice in infection groups were infected with 30 stage III A. cantonensis larvae by gavage, and animals in the control group was given an equal amount of physiological saline. Five mice were collected from each of infection groups on days 7, 14, 21 d and 25 d post-infection, and 5 mice were collected from the control group on the day of oral gavage. The general and focal functional impairment was scored using the Clark scoring method to assess the degree of mouse neurological impairment. Five mice from each of infection groups were sacrificed on days 7, 14, 21 d and 25 d post-infection, and 5 mice from the control group were sacrificed on the day of oral gavage. Mouse brain tissues were sampled, and the pathological changes of brain tissues were dynamically observed using hematoxylin and eosin (HE) staining. Immunofluorescence staining with eosinophilic cationic protein (ECP) and ionized calcium binding adaptor molecule 1 (Iba1) was used to assess the degree of eosinophil infiltration and the counts of microglial cells in mouse brain tissues in each group, and the morphological parameters of microglial cells (skeleton analysis and fractal analysis) were quantified by using Image J software to determine the morphological changes of microglial cells. In addition, the expression of M1 microglia markers Fcγ receptor III (Fcgr3), Fcγ receptor IIb (Fcgr2b) and CD86 antigen (Cd86), M2 microglia markers Arginase 1 (Arg1), macrophage mannose receptor C-type 1 (Mrc1), chitinase-like 3 (Chil3), and phagocytosis genes myeloid cell triggering receptor expressed on myeloid cells 2 (Trem2), CD68 antigen (Cd68), and apolipoprotein E (Apoe) was quantified using real-time quantitative reverse transcription PCR (RT-qPCR) assay in the mouse cerebral cortex of mice post-infection. Results A large number of A. cantonensis larvae were seen on the mouse meninges surface post-infection, and many neuronal nuclei were crumpled and deeply stained, with a large number of bleeding points in the meninges. The median Clark scores of mouse general functional impairment were 0 (interquartile range, 0), 0 (interquartile range, 0.5), 6 (interquartile range, 1.0), 14 (interquartile range, 8.5) points and 20 (interquartile range, 9.0) points in the control group and the 7-d, 14-d, 21-d and 25-d groups, respectively (H = 22.45, P < 0.01), and the median Clark scores of mouse focal functional impairment were 0 (interquartile range, 0), 2 (interquartile range, 2.5), 7 (interquartile range, 3.0), 18 (interquartile range, 5.0) points and 25 (interquartile range, 6.5) points in the control group and the 7-d, 14-d, 21-d and 25-d groups, respectively (H = 22.72, P < 0.01). The mean scores of mice general and focal functional impairment were all higher in the infection groups than in the control group (all P values < 0.05). Immunofluorescence staining showed a significant difference in the eosinophil counts in mouse brain tissues among the five groups (F = 40.05, P < 0.000 1), and the eosinophil counts were significantly higher in mouse brain tissues in the 14-d (3.08 ± 0.78) and 21-d infection groups (5.97 ± 1.37) than in the control group (1.00 ± 0.28) (both P values < 0.05). Semi-quantitative analysis of microglia immunofluorescence showed a significant difference in the counts of microglial cells among the five groups (F = 17.66, P < 0.000 1), and higher Iba1 levels were detected in mouse brain tissues in 14-d (5.75 ± 1.28), 21-d (6.23 ± 1.89) and 25-d infection groups (3.70 ± 1.30) than in the control group (1.00 ± 0.30) (all P values < 0.05). Skeleton and fractal analyses showed that the branch length [(162.04 ± 34.10) μm vs. (395.37 ± 64.11) μm; t = 5.566, P < 0.05] and fractal dimension of microglial cells (1.30 ± 0.01 vs. 1.41 ± 0.03; t = 5.266, P < 0.05) were reduced in mouse brain tissues in the 21-d infection group relative to the control group. In addition, there were significant differences among the 5 groups in terms of M1 and M2 microglia markers Fcgr3 (F = 48.34, P < 0.05), Fcgr2b (F = 55.46, P < 0.05), Cd86 (F = 24.44, P < 0.05), Arg1 (F = 31.18, P < 0.05), Mrc1 (F = 15.42, P < 0.05) and Chil3 (F = 24.41, P < 0.05), as well as phagocytosis markers Trem2 (F = 21.19, P < 0.05), Cd68 (F = 43.95, P < 0.05) and Apoe (F = 7.12, P < 0.05) in mice brain tissues. Conclusions A. cantonensis infections may induce severe pathological injuries in mouse brain tissues that are characterized by massive eosinophil infiltration and persistent activation of microglia cells, thereby resulting in progressive deterioration of neurological functions.

OBJECTIVE: To analyze the carrier rates and profiles of pathogenic and likely pathogenic variants for hearing loss-related genes MYO7A, PCDH15, and CDH23 among neonates in Nanjing city through targeted next-generation sequencing (NGS). METHODS: Heel-prick blood samples were collected from 30 043 newborns delivered at Nanjing Women and Children's Health Care Hospital between March 2022 and April 2024. Dried blood spots were prepared, and genomic DNA was extracted. Targeted NGS was applied to detect variants across the full coding regions of the MYO7A, PCDH15, and CDH23 genes. The carrier rates and profiles of pathogenic and likely pathogenic variants of the three genes were analyzed. This study was approved by the Medical Ethics Committee of Nanjing Maternal and Child Health Care Hospital (Ethics No.: 2021KY-071). RESULTS: The carrier rates of pathogenic and likely pathogenic variants (with ≥ 1 variant site) for the MYO7A, PCDH15, and CDH23 genes were 0.340%, 0.226%, and 0.156%, respectively. A total of 65, 49, and 30 variant types were detected in the MYO7A, PCDH15, and CDH23 genes, respectively. For MYO7A, single base variants were predominant, with the most common variant being c.5581C>T, followed by c.1343+1G>A, c.2837T>G, and c.5660C>T, with allelic frequencies of 0.013% (8/60 086), 0.007% (4/60 086), 0.007% (4/60 086), and 0.007% (4/60 086), respectively. PCDH15 variants were mainly deletions, with the most common variant site being c.4699_4715dupAGAGAAAAGATTCAGAG, followed by c.3441delA, c.440T>G, and c.4733_4736delTCAG, with allelic frequencies of 0.015% (9/60 086), 0.005% (3/60 086), 0.005% (3/60 086), and 0.005% (3/60 086), respectively. For CDH23, single base variants were predominant, with c.6604G>A being the most common, followed by c.6085C>T, c.6050+9G>A, and c.6253+1G>A, with allelic frequencies of 0.013% (8/60 086), 0.012% (7/60 086), 0.005% (3/60 086), and 0.005% (3/60 086). CONCLUSION This study analyzed the carrier rates and profiles of pathogenic and likely pathogenic variants of the MYO7A, PCDH15, and CDH23 genes, which can provide more evidence for the prevention and management of deafness in the region.
Humans ; Cadherins/genetics* ; High-Throughput Nucleotide Sequencing/methods* ; Infant, Newborn ; Female ; Myosin VIIa/genetics* ; Cadherin Related Proteins ; Male ; Hearing Loss/genetics* ; Myosins/genetics* ; Heterozygote

Objective:To analyze the carrier rates and profiles of pathogenic and likely pathogenic variants for hearing loss-related genes MYO7A, PCDH15, and CDH23 among neonates in Nanjing city through targeted next-generation sequencing (NGS). Methods:Heel-prick blood samples were collected from 30 043 newborns delivered at Nanjing Women and Children′s Health Care Hospital between March 2022 and April 2024. Dried blood spots were prepared, and genomic DNA was extracted. Targeted NGS was applied to detect variants across the full coding regions of the MYO7A, PCDH15, and CDH23 genes. The carrier rates and profiles of pathogenic and likely pathogenic variants of the three genes were analyzed. This study was approved by the Medical Ethics Committee of Nanjing Maternal and Child Health Care Hospital (Ethics No.: 2021KY-071). Results:The carrier rates of pathogenic and likely pathogenic variants (with ≥ 1 variant site) for the MYO7A, PCDH15, and CDH23 genes were 0.340%, 0.226%, and 0.156%, respectively. A total of 65, 49, and 30 variant types were detected in the MYO7A, PCDH15, and CDH23 genes, respectively. For MYO7A, single base variants were predominant, with the most common variant being c. 5581C>T, followed by c. 1343+ 1G>A, c. 2837T>G, and c. 5660C>T, with allelic frequencies of 0.013% (8/60 086), 0.007% (4/60 086), 0.007% (4/60 086), and 0.007% (4/60 086), respectively. PCDH15 variants were mainly deletions, with the most common variant site being c. 4699_4715dupAGAGAAAAGATTCAGAG, followed by c. 3441delA, c. 440T>G, and c. 4733_4736delTCAG, with allelic frequencies of 0.015% (9/60 086), 0.005% (3/60 086), 0.005% (3/60 086), and 0.005% (3/60 086), respectively. For CDH23, single base variants were predominant, with c. 6604G>A being the most common, followed by c. 6085C>T, c. 6050+ 9G>A, and c. 6253+ 1G>A, with allelic frequencies of 0.013% (8/60 086), 0.012% (7/60 086), 0.005% (3/60 086), and 0.005% (3/60 086). Conclusion:This study analyzed the carrier rates and profiles of pathogenic and likely pathogenic variants of the MYO7A, PCDH15, and CDH23 genes, which can provide more evidence for the prevention and management of deafness in the region.

Carbapenem-resistant Klebsiella pneumoniae(CRKP),as a superbug pathogen,has become a critical global public health threat,with significantly increased detection rates especially in ICU and immunocompromised patients.Traditional antibiotics are progressively losing efficacy,and CRKP's antibiotic resistance spectrum continues to expand,presenting enormous clinical treatment challenges.Bacteriophages,as viruses specifically infecting bacteria,have emerged as a new therapeutic strategy for alternative or complementary antibiotic treatment due to their strong lytic capabilities,excellent biofilm penetration,low side effects,and ability to synergize with antibiotics.This article systematically reviews the epidemiology and resistance mechanisms of CRKP,principles and mechanisms of bacteriophage therapy,research progress(including in vitro experiments,animal models,and clinical applications),combined therapies,and their advantages and challenges.By incorporating the latest research,the article also proposes future development directions,including constructing genetically engineered bacteriophages,designing"intelligent bacteriophages"through synthetic biology,and establishing clinical regulatory systems.Bacteriophage therapy offers a potential breakthrough in combating CRKP,but its standardization,broad-spectrum applicability,and clinical translation pathways still require in-depth exploration and standardization.

Objective:Objective To analyze the carrier rates and mutational spectrum of pathogenic variants in deafness-associated genes among newborns in Nanjing.Methods:In this cross-sectional study, heel blood samples were collected from 30 043 neonates born at Nanjing Maternity and Child Health Care Hospital between March 2022 and April 2024. DNA was extracted from dried blood spots and subjected to targeted next-generation sequencing of the full coding regions of deafness-associated genes GJB2, SLC26A4, USH2A, MT-RNR1, and MYO15A. Descriptive statistics were used to analyze carrier rates and variant characteristics, with pathogenicity classified according to American College of Medical Genetics and Genomics guidelines. Results:(1) Carrier rates: The overall carrier rate for deafness-associated gene variants was 19.196% (5 767/30 043). GJB2 showed the highest carrier rate at 13.174% (3 958/30 043), followed by SLC26A4 (2.912%, 875/30 043), USH2A (1.524%, 458/30 043), MT- RNR1 (0.959%, 288/30 043), and MYO15A (0.626%, 188/30 043). MT-RNR1 follows mitochondrial inheritance, while others are autosomal recessive. (2) Variant analysis revealed: 25 GJB2 variants with c.109G>A being most frequent (allele frequency 4.925%, 2 959/60 086), followed by c.235delC (1.127%, 677/60 086) and c.299_300delAT (0.261%, 157/60 086); 85 SLC26A4 variants dominated by c.919-2A>G (0.621%, 373/60 086), c.2009T>C (0.165%, 99/60 086), and c.2168A>G (0.100%, 60/60 086); 118 USH2A variants with c.2802T>G (0.218%, 131/60 086) and c.8559-2A>G (0.165%, 99/60 086) most prevalent; three MT-RNR1 variants including m.1095T>C (230 cases), m.1555A>G (52 cases), and m.1494C>T (six cases); and 81 MYO15A variants with c.10250_10252delCCT (0.043%, 26/60 086) being most common. Conclusion:The predominant pathogenic variants in deafness-associated genes among Nanjing neonates are GJB2 c.109G>A and SLC26A4 c.919-2A>G, with MT- RNR1 m.1095T>C representing a significant mitochondrial variant.

Objective To investigate the clinical significance of the combined screening of thyroid stimulating hormone(TSH)and seven candidate pathogenic genes of congenital hypothyroidism(CH)for CH.Methods 16 645 newborns delivered in Nanjing Women and Children's Healthcare Hospital from July 2022 to July 2023 were performed the screening of TSH.Their DNA was extracted from dried blood spots and the chip capture second-generation sequencing technology was used to detect the candidate pathogenic genes,in-cluding dual oxidase 2(DUOX2),dual oxidase maturation factor 2(DUOXA2),prophet of pit-1(PROP1),thyroid-stimulating hor-mone receptor(TSHR),thyroid peroxidase(TPO),thyroglobulin(TG),and paired box 8(PAX8).The sensitivity,specificity,pos-itive predictive value(PPV),and negative predictive value(NPV)of the screening of TSH,candidate genes,and their combination for CH were analyzed.Results A total of 13 CH patients were screened out based on sensitive thyroid stimulating hormone(sTSH)and free thyroxine(FT4),including 3 patients with hyperthyrotropinemia.Among them,11 were screened out by TSH alone,4 were screened out by candidate genes alone,and 2 were screened out by the combination of TSH and candidate genes.The sensitivity,speci-ficity,PPV,and NPV of TSH for screening CH were 84.62％,99.23％,7.91％,and 99.97％,respectively.The sensitivity,specifici-ty,PPV,and NPV of candidate genes for screening CH were 30.77％,99.87％,15.38％,and 99.87％,respectively.The sensitivity,specificity,PPV,and NPV of the combination of TSH and candidate genes for screening CH were 100％,99.09％,7.88％,and 100％,respectively.The primary mutant gene in the samples with positive candidate genes was DUOX2(85.71％),mainly point muta-tions,among which the c.1588A>T variant was the most common(16.67％).PAX8(14.29％)was the second most common variation,and all of the variation point were c.280G>A.No positive samples for the pathogenic variants of DUOXA2,TSHR,PROP1,TPO,and TG were detected.Conclusion The combined screening of TSH and candidate genes helps to improve the screening efficacy of CH.The genetic etiology of CH in Nanjing area may be mainly the variation of DUOX2 and PAX8 genes.

Objective To investigate the clinical significance of the combined screening of thyroid stimulating hormone(TSH)and seven candidate pathogenic genes of congenital hypothyroidism(CH)for CH.Methods 16 645 newborns delivered in Nanjing Women and Children's Healthcare Hospital from July 2022 to July 2023 were performed the screening of TSH.Their DNA was extracted from dried blood spots and the chip capture second-generation sequencing technology was used to detect the candidate pathogenic genes,in-cluding dual oxidase 2(DUOX2),dual oxidase maturation factor 2(DUOXA2),prophet of pit-1(PROP1),thyroid-stimulating hor-mone receptor(TSHR),thyroid peroxidase(TPO),thyroglobulin(TG),and paired box 8(PAX8).The sensitivity,specificity,pos-itive predictive value(PPV),and negative predictive value(NPV)of the screening of TSH,candidate genes,and their combination for CH were analyzed.Results A total of 13 CH patients were screened out based on sensitive thyroid stimulating hormone(sTSH)and free thyroxine(FT4),including 3 patients with hyperthyrotropinemia.Among them,11 were screened out by TSH alone,4 were screened out by candidate genes alone,and 2 were screened out by the combination of TSH and candidate genes.The sensitivity,speci-ficity,PPV,and NPV of TSH for screening CH were 84.62％,99.23％,7.91％,and 99.97％,respectively.The sensitivity,specifici-ty,PPV,and NPV of candidate genes for screening CH were 30.77％,99.87％,15.38％,and 99.87％,respectively.The sensitivity,specificity,PPV,and NPV of the combination of TSH and candidate genes for screening CH were 100％,99.09％,7.88％,and 100％,respectively.The primary mutant gene in the samples with positive candidate genes was DUOX2(85.71％),mainly point muta-tions,among which the c.1588A>T variant was the most common(16.67％).PAX8(14.29％)was the second most common variation,and all of the variation point were c.280G>A.No positive samples for the pathogenic variants of DUOXA2,TSHR,PROP1,TPO,and TG were detected.Conclusion The combined screening of TSH and candidate genes helps to improve the screening efficacy of CH.The genetic etiology of CH in Nanjing area may be mainly the variation of DUOX2 and PAX8 genes.

Objective:To analyze the carrier rates and profiles of pathogenic and likely pathogenic variants for hearing loss-related genes MYO7A, PCDH15, and CDH23 among neonates in Nanjing city through targeted next-generation sequencing (NGS). Methods:Heel-prick blood samples were collected from 30 043 newborns delivered at Nanjing Women and Children′s Health Care Hospital between March 2022 and April 2024. Dried blood spots were prepared, and genomic DNA was extracted. Targeted NGS was applied to detect variants across the full coding regions of the MYO7A, PCDH15, and CDH23 genes. The carrier rates and profiles of pathogenic and likely pathogenic variants of the three genes were analyzed. This study was approved by the Medical Ethics Committee of Nanjing Maternal and Child Health Care Hospital (Ethics No.: 2021KY-071). Results:The carrier rates of pathogenic and likely pathogenic variants (with ≥ 1 variant site) for the MYO7A, PCDH15, and CDH23 genes were 0.340%, 0.226%, and 0.156%, respectively. A total of 65, 49, and 30 variant types were detected in the MYO7A, PCDH15, and CDH23 genes, respectively. For MYO7A, single base variants were predominant, with the most common variant being c. 5581C>T, followed by c. 1343+ 1G>A, c. 2837T>G, and c. 5660C>T, with allelic frequencies of 0.013% (8/60 086), 0.007% (4/60 086), 0.007% (4/60 086), and 0.007% (4/60 086), respectively. PCDH15 variants were mainly deletions, with the most common variant site being c. 4699_4715dupAGAGAAAAGATTCAGAG, followed by c. 3441delA, c. 440T>G, and c. 4733_4736delTCAG, with allelic frequencies of 0.015% (9/60 086), 0.005% (3/60 086), 0.005% (3/60 086), and 0.005% (3/60 086), respectively. For CDH23, single base variants were predominant, with c. 6604G>A being the most common, followed by c. 6085C>T, c. 6050+ 9G>A, and c. 6253+ 1G>A, with allelic frequencies of 0.013% (8/60 086), 0.012% (7/60 086), 0.005% (3/60 086), and 0.005% (3/60 086). Conclusion:This study analyzed the carrier rates and profiles of pathogenic and likely pathogenic variants of the MYO7A, PCDH15, and CDH23 genes, which can provide more evidence for the prevention and management of deafness in the region.

Objective:Objective To analyze the carrier rates and mutational spectrum of pathogenic variants in deafness-associated genes among newborns in Nanjing.Methods:In this cross-sectional study, heel blood samples were collected from 30 043 neonates born at Nanjing Maternity and Child Health Care Hospital between March 2022 and April 2024. DNA was extracted from dried blood spots and subjected to targeted next-generation sequencing of the full coding regions of deafness-associated genes GJB2, SLC26A4, USH2A, MT-RNR1, and MYO15A. Descriptive statistics were used to analyze carrier rates and variant characteristics, with pathogenicity classified according to American College of Medical Genetics and Genomics guidelines. Results:(1) Carrier rates: The overall carrier rate for deafness-associated gene variants was 19.196% (5 767/30 043). GJB2 showed the highest carrier rate at 13.174% (3 958/30 043), followed by SLC26A4 (2.912%, 875/30 043), USH2A (1.524%, 458/30 043), MT- RNR1 (0.959%, 288/30 043), and MYO15A (0.626%, 188/30 043). MT-RNR1 follows mitochondrial inheritance, while others are autosomal recessive. (2) Variant analysis revealed: 25 GJB2 variants with c.109G>A being most frequent (allele frequency 4.925%, 2 959/60 086), followed by c.235delC (1.127%, 677/60 086) and c.299_300delAT (0.261%, 157/60 086); 85 SLC26A4 variants dominated by c.919-2A>G (0.621%, 373/60 086), c.2009T>C (0.165%, 99/60 086), and c.2168A>G (0.100%, 60/60 086); 118 USH2A variants with c.2802T>G (0.218%, 131/60 086) and c.8559-2A>G (0.165%, 99/60 086) most prevalent; three MT-RNR1 variants including m.1095T>C (230 cases), m.1555A>G (52 cases), and m.1494C>T (six cases); and 81 MYO15A variants with c.10250_10252delCCT (0.043%, 26/60 086) being most common. Conclusion:The predominant pathogenic variants in deafness-associated genes among Nanjing neonates are GJB2 c.109G>A and SLC26A4 c.919-2A>G, with MT- RNR1 m.1095T>C representing a significant mitochondrial variant.