Molecular Epidemiological Characteristics Research Articles

Novel Intertypic Recombinant Coxsackievirus A2 Containing Specific Amino Acid Mutations in the RNA-Dependent RNA Polymerase Potentially Associated With Its Emergence.

Coxsackievirus A2 (CVA2), a member of enterovirus A species (EV-A), is associated with diverse human diseases and occasionally causes acute gastroenteritis (AGE). In Thailand, CVA2 emerged as the predominant genotype in 2019. The increasing incidence of CVA2, coupled with the limited availability of full-length genomes, highlights the need for more complete genome sequence analysis to facilitate molecular epidemiology study. This study aimed to investigate the molecular epidemiology, evolutionary dynamics, and recombination characteristics of CVA2 associated with AGE in Thailand from 2013 to 2022. A total of 19 full-genome sequences of CVA2 isolated from stool samples of AGE patients in Thailand were characterized and analyzed together with the reference sequences available in the GenBank database. A novel lineage of CVA2 (subgenotype C5) was detected with the potential recombination with CVA10 within the P2 and P3 regions. Specific consensus amino acid mutations, A61S in the VP3 gene and R136K in the 3D (RdRp) gene, were identified in all CVA2 recombinant strains. Additionally, the S45G mutation in the RdRp gene was found to be potentially associated with the emergence of CVA2 infection in 2019. In conclusion, this study reveals potential intertypic recombinant events and specific mutations in CVA2 strains isolated from AGE patients and provides a broader understanding of its evolutionary epidemiology.

Molecular epidemiology and clinical characteristics of Staphylococcus aureus bacteremia in Japanese adults

IntroductionStaphylococcus aureus bacteremia (SAB), especially when caused by methicillin-resistant S. aureus (MRSA), is of considerable clinical importance. In recent years, the proportion of MRSA among S. aureus has decreased, and a relative increase in the proportion of methicillin-susceptible S. aureus (MSSA) has been observed. It is therefore necessary to consider both MRSA and MSSA when assessing the microbiological and clinical significance of SAB. Materials and MethodsWe included SAB cases from the Nara Medical University Hospital between January 2015 and February 2017. We performed drug susceptibility testing, toxicity gene analysis, multilocus sequence typing (MLST), and polymerase chain reaction-based open reading frame typing (POT) of stored strains to integrate clinical and bacteriological characteristics. ResultsThere were 90 cases during the experimental period (42 MRSA and 48 MSSA), with 30-day mortality rates of 19% for MRSA and 10.4% for MSSA. Deaths were more frequently complicated by septic shock and disseminated intravascular coagulation. MLST studies showed that ST8, ST764, ST1, and ST15 were prevalent in the MRSA group, whereas ST5, ST188, and ST12 were prevalent in MSSA. Infective endocarditis cases had a long time from onset to the initiation of effective antimicrobials and were all MSSA. MLST and POT results correlated well, and POT appeared to have better discriminatory power. ConclusionsThe severity and mortality of SAB, along with the microbiological characteristics of causative isolates, vary by location and time. Continued studies integrating clinical and microbiological investigations are thus needed.

Molecular epidemiology and genetic characterization of hepatitis B virus in two major provinces of Pakistan.

Hepatitis B virus (HBV) infection is a major public health issue and is responsible for considerable morbidity and mortality globally. In Pakistan, the prevalence of chronic HBV infection varies from 2% to 4%, with an estimated exposure rate of approximately 34%. The major objective of this study was to determine the prevalence of HBV genotypes and the pattern of escape mutations in the HBV S gene in two major provinces of Pakistan: Punjab and Khyber Pakhtunkhwa. A total of 146 serum/plasma samples collected from hepatitis B patients were sequenced by the Sanger method. Phylogenetic analysis indicated the intermixed circulation of closely related strains of HBV genotype D and subgenotypes HBV/D1, HBV/D2, and HBV/A2 in both provinces, and various escape mutations (Y100C, P120S/T, G119R, C121S/Y, R122K, T126I, A128V, Q129H/R, G130R, T131N/I, M133T/I, Y134N, C137W, S143L, and D144E) were identified. These findings provide important insights into the current prevalence of HBV genotypes in Pakistan and will help in the establishment of more-efficient disease interventions and patient management strategies.

Virulence and molecular epidemiological analysis of three human blood-borne Streptococcus suis

ObjectiveTo understand the virulence genes and molecular epidemiological characteristics of human-infected strains of Streptococcus suis in Rui'an, Zhejiang Province, from 2021 to 2022, and to provide a scientific basis for diagnosis, treatment, prevention, and control. MethodsThree blood-borne strains of Streptococcus suis were analysed by morphological observation, identification, and drug sensitivity tests. We performed polymerase chain reaction (PCR) amplification of their main seven virulence factors and housekeeping genes. This was followed by virulence analysis and multilocus sequence typing. We analysed their relationships with local pathogens from previous years. ResultsThree Streptococcus suis strains were isolated from the blood samples of three patients. From these, the virulence genotypes demonstrated that the two strains were orf2+ and ef+/orf2+/sly+, respectively. The Multilocus sequence typing (MLST) typing results demonstrated that the two strains were ST25 and ST7, respectively. ConclusionThe first isolation of ST25 Streptococcus suis in Rui'an was presumed to have a close affinity with the endemic strain in North America. The other strain was an ST7 clone, consistent with the endemic strain in Sichuan, and which may have originated from Sichuan. Virulence genotype analysis demonstrated that different virulence genes of the pathogens resulted in different clinical manifestations.

Molecular epidemiological characteristics, variant spectrum and genotype-phenotype correlation of glucose-6-phosphate dehydrogenase deficiency in China: A population-based multicenter study using newborn screening.

Newborn screening (NBS) for glucose-6-phosphate dehydrogenase (G6PD) deficiency by biochemical tests is being used worldwide, however, the outcomes arising from combined genetic and biochemical tests have not been evaluated. This research aimed to evaluate the outcomes of application of combined genetic and biochemical NBS for G6PD deficiency and to investigate the molecular epidemiological characteristics, variant spectrum, and genotype-phenotype correlation of G6PD deficiency in China. A population-based cohort of 29,601 newborns were prospectively recruited from eight NBS centers in China between February 21 and December 30, 2021. Biochemical and genetic NBS was conducted simultaneously. The overall prevalence of G6PD deficiency was 1.12% (1.86% for male, and 0.33% for female; 1.94% for South China and 0.08% for North China). Genetic NBS identified 10 male patients undetected by biochemical NBS. The overall positive predictive values (PPVs) of biochemical and genetic NBS were 79.95% and 47.57%, respectively. A total of 15 variants were identified, with the six most common variants being c.1388G > A, c.1376G > T, c.95A > G, c.871G > A, c.1024C > T and c.392G > T (94.2%). The activity of G6PD was correlated with the type and WHO classification of variants. This study highlighted that combined screening could enhance the efficiency of current NBS for diagnosing G6PD deficiency. The prevalence, variant spectrum and allele frequency of G6PD deficiency vary across different regions. Our data provide valuable references for clinical practice and optimization of future screening strategies for G6PD deficiency.

Genomic epidemiology of ceftriaxone-resistant non-typhoidal Salmonella enterica strain in China

Non-typhoidal Salmonella (NTS) is one of the top causes of diarrhea worldwide. Ceftriaxone is commonly recommended as the initial treatment option for Salmonella infections due to its antibacterial effectiveness. The objective of this study was to investigate the molecular epidemiological characteristics of NTS and to compare the phenotypic and genotypic profiles of antimicrobial resistance in multidrug-resistant Salmonella strains by sequencing 329 NTS strains collected from a county-level hospital between 2018 and 2021. Multi-locus sequence typing (MLST), antimicrobial resistance genes and plasmid types were identified by BacWGSTdb 2.0 webserver. Phylogenetic analysis of all NTS strains was carried out using Snippy and Gubbins software. The transferability of ceftriaxone resistant plasmids was confirmed through plasmid conjugation assays, and verified by S1-PFGE-Southern blot assays. The predominant serotypes among all NTS strains were Typhimurium (161/329), Enteritidis (49/329) and London (45/329). The most common sequence type observed was ST34 (86/329), followed by ST19 (72/329) and ST11 (47/329). The antimicrobial resistance of Salmonella to a wide range of antimicrobials showed an overall increase. Out of these 37 (11.24%) ceftriaxone-resistant strains, with the majority of them (33/37) being blaCTX−M. The predominant plasmid types identified were IncHI2 (14/21) and IncI1 (6/21), ranging in size from 70 kb to 360 kb. The conjugation efficiency was calculated with the high conjugation efficiency of 1.1 × 10− 5 to 9.3 × 10− 2. The strains varied widely, ranging from 3 to 45,024 single nucleotide polymorphisms (SNPs). There are close linkages observed among the predominant lineage, with an average of 78 SNPs between each pair of ST34 strains. The findings contribute to our understanding of the transmission and resistance mechanisms of multidrug-resistant Salmonella, thereby facilitating the development of effective control strategies.

Molecular and Microbiological Characterization of Candida auris Strains Isolated from Colonized and Infected Patients in a New York City Tertiary Care Center

Abstract Candida auris is a multidrug resistant pathogen that causes healthcare associated invasive infections and outbreaks with high mortality rates, often involving intensive care units (ICU). High rates of C. auris colonization among patients in ICUs are associated with increased risk of subsequent C. auris hospital-acquired infections. In this study, microbiological characteristics, and antifungal susceptibility patterns of C. auris isolates collected between January and December 2023 were investigated. Whole-genome sequencing was used to examine the genetic variation and molecular epidemiological characteristics. During the study period, a total of 20 patients were found to be colonized with C. auris by three site skin swab and eight of these patients subsequently developed invasive infection with C. auris. Among patients with both colonization and infection, all invasive isolates were closely related genetically to colonizing isolates and belonged to clade I. Candidemia was the most common infection identified in 4/8 (50%) patients. All isolates were resistant to fluconazole and thirteen isolates from six patients were resistant to amphotericin B. Paired isolates from patients with both colonization and invasive infection showed the same antifungal resistance patterns. All isolates had the ERG11 p.K143R mutation known to cause fluconazole resistance. Three isolates from one patient with a FKS1 hotspot region 3 p.M690I mutation were resistant to caspofungin and two isolates from one patient with a FKS1 hotspot region 2 p.R1354H mutation were resistant to caspofungin, anidulafungin and micafungin. Our study demonstrates that colonization with C. auris is associated with increased risk of infection by the same strain amongst critical ill patients. C. auris screening and implementation of adequate infection control measures are urgently needed to prevent the dissemination of C. auris infections.

Multi-Locus Sequence Typing-Based Genetic Analysis, Antifungal Resistance, and Clinical Prognosis of Cryptococcus neoformans Infections in HIV-Infected Patients in Northern China.

This study aimed to investigate the molecular epidemiological characteristics and drug sensitivity of Cryptococcus from HIV-infected patients and their relationship with patients' prognosis. Seventy-six strains were collected and identified to the species level by matrix-assisted laser desorption ionization-time of flight mass spectrometry, confirmed by internal transcribed spacer sequencing. Multi-locus sequence typing was used for the typing of Cryptococcus, and its antifungal susceptibility was tested using FUNGUS 3. The clinical outcomes of the patients were reviewed at 3-, 6-, 9-, and 12-month follow-ups. All strains were Cryptococcus neoformans var. grubii classified into seven sequence types (STs) dominated by ST5, ST31, and a new ST702 strain. The 6- and 9-month survival rates were highest for patients infected with ST31, ST32, and ST174. The antifungal resistant rates were 13.2%, 2.6%, and 1.4% for fluconazole, amphotericin B, and 5-fluorocytosine. Except itraconazole, the minimum inhibitory concentration (MIC) values and wild type (WT)/non-wild type (NWT) of Cryptococcus for antifungal drugs were not related to the clinical prognosis of HIV-infected patients with cryptococcal infection. ST5 was the main ST type, and the new ST702 type was found in a patient who died in a short period of time. Cryptococcus neoformans var. grubii had a relatively high antifungal drug resistance rate to fluconazole. The WT strain accounted for the highest proportions for 5-fluorocytosine, amphotericin B, fluconazole, voriconazole, and itraconazole. The MIC values of Cryptococcus for first-line antifungal drugs showed no relationship with clinical prognosis, implying that MIC values cannot be used to predict the clinical outcome of these patients.

Characterization of Two Novel HIV-1 CRF01_AE/CRF07_BC Recombinant Forms Among Men Who Have Sex with Men and Mother-to-Child Transmission Cases in Baoding City, China.

CRF01_AE and CRF07_BC are predominant circulating HIV-1 subtypes in China. In this study, we report two novel HIV-1 CRF01_AE/CRF07_BC recombinant forms isolated from one man who has sex with men (MSM) (BDD027) and one mother-to-child transmission (MTCT) case (BDL123) in Baoding City, Hebei Province, China. The recombination breakpoint analysis showed that the recombination pattern of the near-full-length genome of BDD027 consisted of two CRF07_BC fragments inserted into a CRF01_AE backbone, while the recombination pattern of the near-full-length genome of BDL123 consisted of one CRF01_AE fragment inserted into a CRF07_BC backbone. This study demonstrates the importance of strengthening the monitoring of HIV-1 molecular epidemiological characteristics and emphasizes the urgent need to reduce the HIV-1 epidemic among MSM and MTCT populations in China.

Molecular epidemiology and evolutionary characteristics of dengue virus 2 in East Africa

Despite the increasing burden of dengue, the regional emergence of the virus in Kenya has not been examined. This study investigates the genetic structure and regional spread of dengue virus-2 in Kenya. Viral RNA from acutely ill patients in Kenya was enriched and sequenced. Six new dengue-2 genomes were combined with 349 publicly available genomes and phylogenies used to infer gene flow between Kenya and other countries. Analyses indicate two dengue-2 Cosmopolitan genotype lineages circulating in Kenya, linked to recent outbreaks in coastal Kenya and Burkina Faso. Lineages circulating in Western, Southern, and Eastern Africa exhibiting similar evolutionary features are also reported. Phylogeography suggests importation of dengue-2 into Kenya from East and Southeast Asia and bidirectional geneflow. Additional lineages circulating in Africa are also imported from East and Southeast Asia. These findings underscore how intermittent importations from East and Southeast Asia drive dengue-2 circulation in Kenya and Africa more broadly.

The prevalence, antimicrobial resistance and molecular genetic characteristics of fosfomycin-resistant Klebsiella pneumoniae isolated from raw milk

This investigation aims to explore the molecular epidemiological characteristics of the fosfomycin resistance gene, fosA5, in unpasteurized raw milk. A total of 385 unpasteurized raw milk samples were collected from dairy farms in Jiangsu province, China. From these samples, 102 strains of Klebsiella pneumoniae were isolated, with 9 of them testing positive for the presence of the fosA5 gene. Antimicrobial susceptibility testing revealed that K. pneumoniae carrying fosA5 exhibited high-level resistance to fosfomycin but also to other tested antibiotics, indicating multidrug resistance. The S1-PFGE and conjugation results indicated that the fosA5 gene was located on the chromosome. Furthermore, the analysis of PFGE patterns, MLST, and WGS elucidated the genetic diversity among K. pneumoniae strains harboring the fosA5 gene, indicating both clonal and horizontal transmission. This study highlights the prevalence and characteristics of the fosA5 fosfomycin resistance gene in large-scale dairy farms in Jiangsu province. Given its potential public health risks, these findings warrant significant attention.

Retrospective analysis of molecular characteristics, risk factors, and outcomes in carbapenem-resistant Klebsiella pneumoniae bloodstream infections

BackgroundKlebsiella pneumoniae (KP) is the second most prevalent Gram-negative bacterium causing bloodstream infections (BSIs). In recent years, the management of BSIs caused by KP has become increasingly complex due to the emergence of carbapenem-resistant Klebsiella pneumoniae (CRKP). Although numerous studies have explored the risk factors for the development of CRKP-BSIs, the mortality of patients with KP-BSIs, and the molecular epidemiological characteristics of CRKP, the variability in data across different populations, countries, and hospitals has led to inconsistent conclusions. In this single-center retrospective observational study, we utilized logistic regression analyses to identify independent risk factors for CRKP-BSIs and factors associated with mortality in KP-BSI patients. Furthermore, a risk factor-based prediction model was developed. CRKP isolates underwent whole-genome sequencing (WGS), followed by an evaluation of microbiological characteristics, including antimicrobial resistance and virulence genes, as well as epidemiological characteristics and phylogenetic analysis.ResultsOur study included a total of 134 patients with KP-BSIs, comprising 50 individuals infected with CRKP and 84 with carbapenem-susceptible Klebsiella pneumoniae (CSKP). The independent risk factors for CRKP-BSIs were identified as gastric catheterization (OR = 9.143; CI = 1.357–61.618; P = 0.023), prior ICU hospitalization (OR = 4.642; CI = 1.312–16.422; P = 0.017), and detection of CRKP in non-blood sites (OR = 8.112; CI = 2.130-30.894; P = 0.002). Multivariate analysis revealed that microbiologic eradication after 6 days (OR = 3.569; CI = 1.119–11.387; P = 0.032), high Pitt bacteremia score (OR = 1.609; CI = 1.226–2.111; P = 0.001), and inappropriate empirical treatment after BSIs (OR = 6.756; CI = 1.922–23.753; P = 0.003) were independent risk factors for the 28-day mortality in KP-BSIs. The prediction model confirmed that microbiologic eradication after 6.5 days and a Pitt bacteremia score of 4.5 or higher were significant predictors of the 28-day mortality. Bioinformatics analysis identified ST11 as the predominant CRKP sequence type, with blaKPC−2 as the most prevalent gene variant. CRKP stains carried multiple plasmid-mediated resistance genes along with some virulence genes. Phylogenetic analysis indicated the presence of nosocomial transmission of ST11 CRKP within the ICU.ConclusionsThe analysis of risk factors for developing CRKP-BSIs and the association between KP-BSIs and 28-day mortality, along with the development of a risk factor-based prediction model and the characterization of CRKP strains, enhances clinicians’ understanding of the pathogens responsible for BSIs. This understanding may help in the timely administration of antibiotic therapy for patients with suspected KP-BSIs, potentially improving outcomes.

The biological characteristics of DAstV molecular epidemiology and pathogenicity of duck astrovirus causing hepatitis in ducks and chickens in Southeast China

A suspected outbreak of duck astrovirus (DAstV) disease occurred in a large Muscovy duck farm in Guangdong Province, China, in June 2022, which severely affected the production performance and health of Muscovy ducks. This study aimed to investigate the prevalence of DAstV disease in Southeast China. Herein, we employed semi-nested PCR ethodto screen 5203 swab and liver samples from 11 Muscovy duck farms in 5 provinces of China for the presence of DAstV. Among them, 1356 samples (26.06%, 1356/5203) tested positive for DAstV, out of which 11 DAstV strains were isolated after 10 generations of blind transmission through Leghorn male hepatoma (LMH) cells and performed their whole-genome sequencing. The alignment results showed that the 11 DAstV isolates exhibited relatively low homology (15.4%-75%) with the astrovirus isolates from other species published in GenBank, whereas their homology (nucleotide: 90.4%-99.99%; amino acid: 94%-99.8%) with the DAstV type 1 (DAstV-1) reference strain was higher, indicating considerable homology. The results indicated that DAstV-1 was the main pathogenic factor. Herein, we successfully recreated the clinical symptoms of natural infection in 28-day-old specific-pathogen-free (SPF) ducks using the DAstV-1-GDB-2022 strain. The primary clinical manifestations included liver enlargement, hemorrhaging, and disruptions in liver function. Additionally, we confirmed the cross-species transmission potential of DAstV-1, marking the first occurrence of clinical symptoms of DAstV in 28-day-old SPF chickens. Our findings provide new perspectives on the epidemiology and pathogenicity of DAstV-1 and may help in advancing the development of DAstV vaccines.

Epidemiological characteristics of human metapneumovirus among children in Nanjing, China.

The objective of this study was to examine the molecular epidemiology and clinical characteristics of HMPV infection among children with ARIs in Nanjing. The respiratory samples were collected from 2078 children (≤ 14 years) with acute respiratory infections and were tested for HMPV using real-time RT-PCR. Amplification and sequencing of the HMPV G gene were followed by phylogenetic analysis using MEGA 7.0. The detection rate of HMPV among children was 4.7% (97/2078), with a concentration in those under 5 years of age. Notably, the peak season for HMPV prevalence was observed in winter. Among the 97 HMPV-positive samples, 51.5% (50/97) were available for characterization of the HMPV G protein gene. Phylogenetic analysis indicated that the sequenced HMPV strains were classified into three sublineages: A2c111nt - dup (84.0%), B1 (2.0%), and B2 (14.0%). There was an incidence of HMPV among hospitalized children during 2021-2022 in Nanjing with A2c111nt - dup being the dominant strain. This study demonstrated the molecular epidemiological characteristics of HMPV among children with respiratory infections in Nanjing, China.

The novel 2024 WHO Neisseria gonorrhoeae reference strains for global quality assurance of laboratory investigations and superseded WHO N. gonorrhoeae reference strains-phenotypic, genetic and reference genome characterization.

MDR and XDR Neisseria gonorrhoeae strains remain major public health concerns internationally, and quality-assured global gonococcal antimicrobial resistance (AMR) surveillance is imperative. The WHO global Gonococcal Antimicrobial Surveillance Programme (GASP) and WHO Enhanced GASP (EGASP), including metadata and WGS, are expanding internationally. We present the phenotypic, genetic and reference genome characteristics of the 2024 WHO gonococcal reference strains (n = 15) for quality assurance worldwide. All superseded WHO gonococcal reference strains (n = 14) were identically characterized. The 2024 WHO reference strains include 11 of the 2016 WHO reference strains, which were further characterized, and four novel strains. The superseded WHO reference strains include 11 WHO reference strains previously unpublished. All strains were characterized phenotypically and genomically (single-molecule PacBio or Oxford Nanopore and Illumina sequencing). The 2024 WHO reference strains represent all available susceptible and resistant phenotypes and genotypes for antimicrobials currently and previously used (n = 22), or considered for future use (n = 3) in gonorrhoea treatment. The novel WHO strains include internationally spreading ceftriaxone resistance, ceftriaxone resistance due to new penA mutations, ceftriaxone plus high-level azithromycin resistance and azithromycin resistance due to mosaic MtrRCDE efflux pump. AMR, serogroup, prolyliminopeptidase, genetic AMR determinants, plasmid types, molecular epidemiological types and reference genome characteristics are presented for all strains. The 2024 WHO gonococcal reference strains are recommended for internal and external quality assurance in laboratory examinations, especially in the WHO GASP, EGASP and other GASPs, but also in phenotypic and molecular diagnostics, AMR prediction, pharmacodynamics, epidemiology, research and as complete reference genomes in WGS analysis.

Molecular Epidemiological Characteristics of Carbapenem Resistant Aeromonas from Hospital Wastewater.

Hospital wastewater (HWW) promotes the spread of carbapenem resistance genes (CRGs). Aeromonas carry a large number of CRGs in HWW, they may play a role as a suitable reservoir for CRGs, while resistomes in HWW are still poorly characterized regarding carbapenem resistant Aeromonas. Thus, the aim of the study was to evaluate the molecular epidemiological characteristics of carbapenem resistant Aeromonas in HWW. A total of 33 carbapenem resistant Aeromonas were isolated from HWW. Antimicrobial susceptibility testing and polymerase chain reaction (PCR) were used to assess the antimicrobial resistance profiles. Molecular typing was performed using enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) and multilocus sequence typing (MLST). The horizontal transmission mode of bla KPC was explored through conjugation and transformation experiments. The stability of bla KPC-IncP-6 plasmids was assessed through plasmid stability and in vitro competition test. The PCR mapping method was used to investigate the structural diversity of bla KPC. The detection rates of bla KPC and cphA in Aeromonas were 97.0% and 39.4% respectively. Aeromonas caviae were grouped into 13 clusters by ERIC-PCR and 12 STs by MLST. Aeromonas veronii were grouped into 11 clusters by ERIC-PCR and 4 STs by MLST. 56.3% bla KPC were located on mobilizable IncP-6 plasmids. bla KPC-IncP-6 plasmid showed high stability and low cost fitness. Carbapenem resistant Aeromonas from HWW mainly carried bla KPC, which exhibited great structural diversity. Aeromonas might serve as reservoirs for bla KPC and bla KPC might spread mainly through transformation in HWW.

Virulence gene distribution and molecular epidemiological characteristics of carbapenem-resistant Klebsiella pneumoniae in the ICU.

The drug-resistant genes carried by carbapenem-resistant Klebsiella pneumoniae (CRKP) limit clinical treatment options, and its virulence genes severely affect patient prognosis. This study aims to investigate the distribution of virulence genes, capsular serotypes, and molecular epidemiological characteristics of CRKP in ICU, to understand the characteristics of CRKP infections in ICU, and to provide a scientific basis for effective monitoring and control of CRKP infections in ICU. A total of 40 non-duplicate strains of CRKP isolated from the ICU of Guangdong Provincial People's Hospital between January 2021 and December 2022 were collected and analyzed. Whole-genome sequencing was used to analyze the distribution of resistance genes, virulence genes, and capsular serotypes of the strains. The sequences of 7 housekeeping genes of CRKP genome were uploaded to the Klebsiella pneumoniae (KPN)multilocus sequence typing (MLST) database to determine the sequence types (STs) of the strains. The age of the 40 ICU CRKP-infected patients was (69.03±17.82) years old, with various underlying diseases, and there were 20 patients with improved clinical outcome and 20 patients with death. The isolated strains primarily originated from mid-stream urine and bronchoalveolar lavage fluid. Whole-genome sequencing results revealed that the strains predominantly carried blaKPC-1 (29 strains, 72.5%) and blaNDM-1 (6 strains, 15.0%), with 5 strains carrying both blaKPC-1 and blaNDM-1. Various virulence genes were detected, among which the carriage rates of genes such as entA, entB, entE, entS, fepA, fepC, fepG, yag/ecp, and ompA reached 100%, while the carriage rates of genes such as entD, fimB, iroB, iroD, fes,and pla were low. The CRKP strains isolated from ICU were predominantly ST11 (27 cases, 67.5%), with KL64 being the main capsular serotype (29 cases, 72.5%). A total of 23 ST11-KL64 CRKP strains were detected, accounting for 57.5%. The main type of ICU CRKP is ST11-KL64, carrying various virulence genes, primarily those related to iron absorption. Furthermore, blaKPC has shifted from blaKPC-2 to blaKPC-1. Therefore, close monitoring of the molecular epidemiological changes of CRKP is necessary, and strict control measures should be implemented to effectively curb the occurrence of CRKP infections.

First report of molecular epidemiology and phylogenetic characteristics of feline herpesvirus (FHV-1) from naturally infected cats in Kunshan, China

BackgroundFeline herpesvirus type 1 (FHV-1) is a life threatening highly contagious virus in cats and typically causes upper respiratory tract infections as well as conjunctival and corneal ulcers. Genetic variability could alter the severity of diseases and clinical signs. Despite regular vaccine practices against FHV-1 in China, new FHV-1 cases still commonly occur. The genetic and phylogenetic characteristics of FHV-1 in Kunshan city of China has not been studied yet. Therefore, this study was planned to investigate the prevalence, molecular characteristics of circulating strains, and phylogenetic analyses of FHV-1. This is the first report of molecular epidemiology and phylogenetic characteristics of FHV-1 from naturally infected cats in Kunshan, China.MethodsThe occulo-nasal swabs were collected from diseased cats showing respiratory distress, conjunctivitis, and corneal ulcers at different veterinary clinics in Kunshan from 2022 to 2023. Clinical data and general information were recorded. Swab samples were processed for preliminary detection of FHV-1. Thymidine kinase (TK), glycoprotein B (gB) and glycoprotein D (gD) genes were sequenced and analyzed to investigate genetic diversity and evolution of FHV-1.ResultsThe FHV-1 genome was detected in 43 (43/200, 21.5%) samples using RT-PCR targeting the TK gene. Statistical analysis showed a significant correlation between age, vaccination status and living environment (p < 0.05) with FHV-1 positivity, while a non-significant correlation was observed for FHV-1 positivity and sex of cats (p > 0.05). Additionally, eight FHV-1 positive cats were co-infected with feline calicivirus (8/43,18.6%). FHV-1 identified in the present study was confirmed as FHV-1 based on phylogenetic analyses. The sequence analyses revealed that 43 FHV-1 strains identified in the present study did not differ much with reference strains within China and worldwide. A nucleotide homology of 99-100% was determined among gB, TK and gD genes nucleotide sequences when compared with standard strain C-27 and vaccine strains. Amino acid analysis showed some amino acid substitutions in TK, gB and gD protein sequences. A potential N-linked glycosylation site was observed in all TK protein sequences. Phylogenetic analyses revealed minor variations and short evolutionary distance among FHV-1 strains detected in this study.ConclusionsOur findings indicate that genomes of 43 FHV-1 strains are highly homogenous and antigenically similar, and the degree of variation in major envelope proteins between strains is low. This study demonstrated some useful data about prevalence, genetic characteristics, and evolution of FHV-1 in Kunshan, which may aid in future vaccine development.

Clonal transmission of blaIMP-4-carrying ST196 Klebsiella pneumoniae isolates mediated by the IncN plasmid in China

ObjectivesTo investigate the clinical and molecular epidemiological characteristics of blaIMP-4-carrying Klebsiella pneumoniae in a tertiary hospital in China. MethodsTen carbapenem-resistant K. pneumoniae (CRKP) isolates carrying the blaIMP-4 gene were collected. Molecular characteristics were analysed using whole-genome sequencing. Plasmid conjugation experiments were used to analyse conjugation of the plasmids. We compared and analysed K. pneumoniae-carrying blaIMP-4 genomic datasets obtained from the National Center for Biotechnology Information (NCBI) with the strains in this study. ResultsAll 10 CRKP isolates carrying blaIMP-4 were collected from 10 adult patients in the respiratory intensive care unit. These strains were only sensitive to polymyxins and tigecycline due to them simultaneously carrying multiple resistance genes, namely blaOKP-A-5, fosA, oqxA, and oqxB. Notably, R29 harboured two carbapenemase genes (blaNDM-1 and blaIMP-4). These strains had similar drug-resistant phenotypes and genes, all belonging to sequence type (ST)196. Additionally, the patients had experienced spatiotemporal intersection during hospitalization, suggesting that these strains underwent clonal transmission, but they belonged to different clonal clusters from the blaIMP-4-positive K. pneumoniae currently published in the NCBI. Among the 10 strains, blaIMP-4 was located on the IncN plasmid, and six strains had successfully transferred the plasmid to the recipient strain EC600 through plasmid conjugation. ConclusionsThe blaIMP-4-positive ST196 CRKP isolate showed clonal distribution in the respiratory intensive care unit, which was mediated by the IncN plasmid. Consequently, there should be increased monitoring of carbapenem-resistant strains in clinical settings to prevent and control its transmission.

Epidemiological and clinical characterization of community, healthcare-associated and nosocomial colonization and infection due to carbapenemase-producing Klebsiella pneumoniae and Escherichia coli in Spain.

Community-acquired (CA) and healthcare-associated (HCA) infections caused by carbapenemase-producing Enterobacterales (CPE) are not well characterized. The objective was to provide detailed information about the clinical and molecular epidemiological features of nosocomial, HCA and CA infections caused by carbapenemase-producing Klebsiella pneumoniae (CP-Kp) and Escherichia coli (CP-Ec). A prospective cohort study was performed in 59 Spanish hospitals from February to March 2019, including the first 10 consecutive patients from whom CP-Kp or CP-Ec were isolated. Patients were stratified according to acquisition type. A multivariate analysis was performed to identify the impact of acquisition type in 30-day mortality. Overall, 386 patients were included (363 [94%] with CP-Kp and 23 [6%] CP-Ec); in 296 patients (76.3%), the CPE was causing an infection. Acquisition was CA in 31 (8.0%) patients, HCA in 183 (47.4%) and nosocomial in 172 (48.3%). Among patients with a HCA acquisition, 100 (54.6%) had been previously admitted to hospital and 71 (38.8%) were nursing home residents. Urinary tract infections accounted for 19/23 (82.6%), 89/130 (68.5%) and 42/143 (29.4%) of CA, HCA and nosocomial infections, respectively. Overall, 68 infections (23%) were bacteremia (8.7%, 17.7% and 30.1% of CA, HCA and nosocomial, respectively). Mortality in infections was 28% (13%, 14.6% and 42.7% of CA, HCA and nosocomial, respectively). Nosocomial bloodstream infections were associated with increased odds for mortality (adjusted OR, 4.00; 95%CI 1.21-13.19). HCA and CA infections caused by CPE are frequent and clinically significant. This information may be useful for a better understanding of the epidemiology of CPE.