Pub Date : 2025-10-15DOI: 10.1016/j.ijmm.2025.151679
Shereen O. Abd Algaffar , Annelies Verbon , Kimberly Eadie , Deborah Horst-Kreft , Sami A. Khalid , Wendy W.J. van de Sande
Mycetoma is a neglected tropical disease characterized by large tumorous lesions. It can be caused by fungi (eumycetoma) or bacteria (actinomycetoma). The hallmark of mycetoma is the formation of grains by the causative agent. Grains can only be formed in vivo; therefore, in vivo models are crucial to studying mycetoma. In vivo, grain models have been developed in the invertebrate Galleria mellonella for eumycetoma, but not for actinomycetoma. Here, we aimed to develop the first actinomycetoma grain model in G. mellonella larvae. Actinomadura madurae strains DSM43236 and DSM44005 were used to infect G. mellonella larvae. Larval survival was monitored over 10 days. Grain formation was studied histologically and compared to grains in human tissues. The efficacy of trimethoprim-sulfamethoxazole and amikacin, the current standard treatment for actinomycetoma, was determined. A. madurae infection decreased the survival of G. mellonella larvae in a concentration-dependent manner. Grains were formed within 24 h post-infection. After 72 h these grains became melanised. No significantly enhanced survival was noted with trimethoprim-sulfamethoxazole, amikacin, or a combination thereof. In this model, the melanised A. madurae grains did differ from human grains, most likely due to the immune system of G. mellonella. The lack of therapeutic efficacy could be caused by this melanin or the fact that A. madurae grains, in general, are less susceptible to these drugs. More research will be needed to address this question.
{"title":"The development of an Actinomadura madurae grain model in Galleria mellonella larvae","authors":"Shereen O. Abd Algaffar , Annelies Verbon , Kimberly Eadie , Deborah Horst-Kreft , Sami A. Khalid , Wendy W.J. van de Sande","doi":"10.1016/j.ijmm.2025.151679","DOIUrl":"10.1016/j.ijmm.2025.151679","url":null,"abstract":"<div><div>Mycetoma is a neglected tropical disease characterized by large tumorous lesions. It can be caused by fungi (eumycetoma) or bacteria (actinomycetoma). The hallmark of mycetoma is the formation of grains by the causative agent. Grains can only be formed <em>in vivo;</em> therefore, <em>in vivo</em> models are crucial to studying mycetoma<em>. In vivo,</em> grain models have been developed in the invertebrate <em>Galleria mellonella</em> for eumycetoma, but not for actinomycetoma. Here, we aimed to develop the first actinomycetoma grain model in <em>G. mellonella</em> larvae. <em>Actinomadura madurae</em> strains DSM43236 and DSM44005 were used to infect <em>G. mellonella</em> larvae. Larval survival was monitored over 10 days. Grain formation was studied histologically and compared to grains in human tissues. The efficacy of trimethoprim-sulfamethoxazole and amikacin, the current standard treatment for actinomycetoma, was determined. <em>A. madurae</em> infection decreased the survival of <em>G. mellonella</em> larvae in a concentration-dependent manner. Grains were formed within 24 h post-infection. After 72 h these grains became melanised. No significantly enhanced survival was noted with trimethoprim-sulfamethoxazole, amikacin, or a combination thereof. In this model, the melanised <em>A. madurae</em> grains did differ from human grains, most likely due to the immune system of <em>G. mellonella</em>. The lack of therapeutic efficacy could be caused by this melanin or the fact that <em>A. madurae</em> grains, in general, are less susceptible to these drugs. More research will be needed to address this question.</div></div>","PeriodicalId":50312,"journal":{"name":"International Journal of Medical Microbiology","volume":"321 ","pages":"Article 151679"},"PeriodicalIF":3.6,"publicationDate":"2025-10-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145318953","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-09-23DOI: 10.1016/j.ijmm.2025.151676
Leonie H. Aldejohann , Joerg Steinmann , Tamara Ruegamer , Ronny Martin , Nadja Thielemann , Grit Walther , Oliver Kurzai , Alexander M. Aldejohann
Background
COVID-19-associated-pulmonary-aspergillosis (CAPA) is a severe superinfection mostly affecting critically ill COVID-19 patients. Early diagnosis and clinical management of CAPA remain major clinical challenges.
Here, we evaluated different approaches to classify culture-positive CAPA at its peak season, assessed incidence and mortality, identified risk factors and analysed clinical and laboratory CAPA-management of three German tertiary care hospitals.
Methods
A retrospective multi-center analysis was performed. Inclusion criteria were SARS-CoV-2-positivity, Aspergillus-culture-positivity of lower respiratory tract specimen and ARDS. Cases were primarily classified according to ECMM/ISHAM-criteria. Species-ID was confirmed by each center. Susceptibility was assessed by EUCAST-microdilution or VIPcheck-screening. Statistical analysis revealed mortality affecting factors.
Results
95 culture-positive CAPA cases were classified as possible (36/95) or probable (59/95) by ECMM/ISHAM; 54 probable cases matched 2 or 3 additional classifications. Incidence rates were higher in ICU (2020/21: 1.56 %/2.13 % non-ICU vs. 5.14 %/6.77 % ICU). A. fumigatus was the most abundant species (93 %; (88/95)). Most patients received steroids to treat COVID-19-ARDS and required respiratory support (steroids: 71 % (67/95); intubated patients 52 % (49/95); ECMO (48 % (46/95)). Retrospective evaluation showed adherence to ECMM/ISHAM antifungal therapy guideline in 71 % (67/95). Case fatality rate was 60 % (57/95). A significant association between GM indices > 3 in respiratory fluid or nicotine abuse (p = 0.035 FE, OR=0.252, 95 % CI=0.066–0.986) and mortality was observed in univariate analysis. Convalescent plasma therapy was significantly associated with mortality reduction in uni- and multivariate analysis (p = 0.020).
Conclusion
Our data reveal regional differences in prevalence, diagnosis, and treatment of culture-positive CAPA in Germany. We could identify new factors affecting survival or mortality.
{"title":"Culture-positive COVID-19-associated pulmonary aspergillosis (CAPA) in Germany","authors":"Leonie H. Aldejohann , Joerg Steinmann , Tamara Ruegamer , Ronny Martin , Nadja Thielemann , Grit Walther , Oliver Kurzai , Alexander M. Aldejohann","doi":"10.1016/j.ijmm.2025.151676","DOIUrl":"10.1016/j.ijmm.2025.151676","url":null,"abstract":"<div><h3>Background</h3><div>COVID-19-associated-pulmonary-aspergillosis (CAPA) is a severe superinfection mostly affecting critically ill COVID-19 patients. Early diagnosis and clinical management of CAPA remain major clinical challenges.</div><div>Here, we evaluated different approaches to classify culture-positive CAPA at its peak season, assessed incidence and mortality, identified risk factors and analysed clinical and laboratory CAPA-management of three German tertiary care hospitals.</div></div><div><h3>Methods</h3><div>A retrospective multi-center analysis was performed. Inclusion criteria were SARS-CoV-2-positivity, <em>Aspergillus</em>-culture-positivity of lower respiratory tract specimen and ARDS. Cases were primarily classified according to ECMM/ISHAM-criteria. Species-ID was confirmed by each center. Susceptibility was assessed by EUCAST-microdilution or VIPcheck-screening. Statistical analysis revealed mortality affecting factors.</div></div><div><h3>Results</h3><div>95 culture-positive CAPA cases were classified as possible (36/95) or probable (59/95) by ECMM/ISHAM; 54 probable cases matched 2 or 3 additional classifications. Incidence rates were higher in ICU (2020/21: 1.56 %/2.13 % non-ICU vs. 5.14 %/6.77 % ICU). <em>A. fumigatus</em> was the most abundant species (93 %; (88/95)). Most patients received steroids to treat COVID-19-ARDS and required respiratory support (steroids: 71 % (67/95); intubated patients 52 % (49/95); ECMO (48 % (46/95)). Retrospective evaluation showed adherence to ECMM/ISHAM antifungal therapy guideline in 71 % (67/95). Case fatality rate was 60 % (57/95). A significant association between GM indices > 3 in respiratory fluid or nicotine abuse (p = 0.035 FE, OR=0.252, 95 % CI=0.066–0.986) and mortality was observed in univariate analysis. Convalescent plasma therapy was significantly associated with mortality reduction in uni- and multivariate analysis (p = 0.020).</div></div><div><h3>Conclusion</h3><div>Our data reveal regional differences in prevalence, diagnosis, and treatment of culture-positive CAPA in Germany. We could identify new factors affecting survival or mortality.</div></div>","PeriodicalId":50312,"journal":{"name":"International Journal of Medical Microbiology","volume":"321 ","pages":"Article 151676"},"PeriodicalIF":3.6,"publicationDate":"2025-09-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145158884","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-09-21DOI: 10.1016/j.ijmm.2025.151675
Deng Hui , Zhu Bin , Zhang Shiyu , Zhang Bin , Dilihumar Zaire , Gao Ruihan , Liu Shuting , Zhou Xin , Zhou Shunchang , Xiong Jian , Yang Xuecheng , Feng Xuemei , Lu Yinping , Zheng Xin , Wang Baoju
Hepatitis B virus (HBV) infection seems to be related to gut microbiota. This study aims to explore the effects of HBV infection on gut microbiota and possible immunological mechanisms using the Chinese woodchuck model. Nine adult woodchucks were randomly divided into Cyclosporine A (CsA) or Control group. Animals were orally treated with CsA and saline for 2 weeks before WHV inoculation, and continued until 8 weeks after that. Blood CsA concentrations were tested at 2 weeks after administration and before discontinuation. Quantitative PCR was used to detect serum WHV DNA. Flow cytometry was used to detect T cell immune response. Feces were collected for 16S rRNA sequencing. The result shows CsA oral administration can reach effective blood drug concentration in woodchucks and successfully prolong WHV replication. After 2 weeks of oral treatment, there was no significant difference in the gut microbiota between the two groups. At the clearance period of serum WHV, the relative abundance of Prevotella and Prevotella genera in the phylum Bacteroidetes significantly increased, while the relative abundance of Firmicutes significantly decreased. Meanwhile, the CD107a degranulation of CD4-T cells in peripheral blood showed a decreasing trend, while there was no significant difference in the frequency of PD-1+ CD4-T cells. In Conclusion, oral administration of CsA can significantly prolong the replication time of WHV in Chinese woodchucks. The gut microbiota of Chinese woodchuck undergoes significant changes during serum WHV clearance, which implies the Chinese woodchuck model can be used to study the interaction between HBV infection and gut microbiota.
{"title":"The impact of HBV infection on gut microbiota using Chinese woodchuck model with woodchuck hepatitis virus (WHV) infection","authors":"Deng Hui , Zhu Bin , Zhang Shiyu , Zhang Bin , Dilihumar Zaire , Gao Ruihan , Liu Shuting , Zhou Xin , Zhou Shunchang , Xiong Jian , Yang Xuecheng , Feng Xuemei , Lu Yinping , Zheng Xin , Wang Baoju","doi":"10.1016/j.ijmm.2025.151675","DOIUrl":"10.1016/j.ijmm.2025.151675","url":null,"abstract":"<div><div>Hepatitis B virus (HBV) infection seems to be related to gut microbiota. This study aims to explore the effects of HBV infection on gut microbiota and possible immunological mechanisms using the Chinese woodchuck model. Nine adult woodchucks were randomly divided into Cyclosporine A (CsA) or Control group. Animals were orally treated with CsA and saline for 2 weeks before WHV inoculation, and continued until 8 weeks after that. Blood CsA concentrations were tested at 2 weeks after administration and before discontinuation. Quantitative PCR was used to detect serum WHV DNA. Flow cytometry was used to detect T cell immune response. Feces were collected for 16S rRNA sequencing. The result shows CsA oral administration can reach effective blood drug concentration in woodchucks and successfully prolong WHV replication. After 2 weeks of oral treatment, there was no significant difference in the gut microbiota between the two groups. At the clearance period of serum WHV, the relative abundance of Prevotella and Prevotella genera in the phylum Bacteroidetes significantly increased, while the relative abundance of Firmicutes significantly decreased. Meanwhile, the CD107a degranulation of CD4-T cells in peripheral blood showed a decreasing trend, while there was no significant difference in the frequency of PD-1+ CD4-T cells. In Conclusion, oral administration of CsA can significantly prolong the replication time of WHV in Chinese woodchucks. The gut microbiota of Chinese woodchuck undergoes significant changes during serum WHV clearance, which implies the Chinese woodchuck model can be used to study the interaction between HBV infection and gut microbiota.</div></div>","PeriodicalId":50312,"journal":{"name":"International Journal of Medical Microbiology","volume":"321 ","pages":"Article 151675"},"PeriodicalIF":3.6,"publicationDate":"2025-09-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145119465","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-09-20DOI: 10.1016/j.ijmm.2025.151678
Cheng-Yen Kao , Nevia Longjam , Jazon Harl Hidrosollo , Lee-Chung Lin , Jang-Jih Lu
Staphylococcus lugdunensis, an emerging coagulase-negative Staphylococcus (CoNS) pathogen, has garnered increasing interest due to its production of lugdunin, a thiazolidine-containing antimicrobial peptide. However, standardized protocols for directly assessing lugdunin activity produced by S. lugdunensis remain lacking. In this study, we examined the effects of pH and incubation duration on lugdunin activity and evaluated the antibacterial spectrum of lugdunin produced by S. lugdunensis isolates against a panel of gram-positive and gram-negative bacterial strains. The optimal conditions for lugdunin antibacterial activity of isolate CGMH-SL85 were identified as pH 7.5 and a 72-h incubation period. Under the tested conditions, the lugdunin produced by CGMH-SL85 exhibited antimicrobial activity against five gram-positive strains, including Staphylococcus aureus ATCC29213 and Staphylococcus haemolyticus CGMH-SH53, followed by Enterococcus faecium EF029 and EF081–2 and Listeria monocytogenes ATCC10403S. However, no antibacterial activity was observed against any of the 11 tested gram-negative bacterial species. Furthermore, four distinct lugdunin susceptibility phenotypes were observed among 47 lugdunin-nonproducing S. lugdunensis strains (14 sequence type (ST)4, 27 ST27, and 6 ST29 strains), including Type A characterized by large, clear inhibition zones; Type B with smaller, clear zones; Type C displaying halo-like inhibition zones; and Type D showing no detectable activity. Moreover, 20 S. lugdunensis strains (42.6 %) exhibited the Type C phenotype. Notably, all six ST29 strains displayed the Type C phenotype, while the Type A phenotype was observed only among ST27 strains (3 strains). In conclusion, we developed a standardized protocol for evaluating lugdunin activity, using pH 7.5 and a 72-h incubation period, and found that different S. lugdunensis strains exhibited distinct lugdunin susceptibility phenotypes.
{"title":"Optimization and spectrum characterization of the antibacterial activity of lugdunin","authors":"Cheng-Yen Kao , Nevia Longjam , Jazon Harl Hidrosollo , Lee-Chung Lin , Jang-Jih Lu","doi":"10.1016/j.ijmm.2025.151678","DOIUrl":"10.1016/j.ijmm.2025.151678","url":null,"abstract":"<div><div><em>Staphylococcus lugdunensis</em>, an emerging coagulase-negative <em>Staphylococcus</em> (CoNS) pathogen, has garnered increasing interest due to its production of lugdunin, a thiazolidine-containing antimicrobial peptide. However, standardized protocols for directly assessing lugdunin activity produced by <em>S. lugdunensis</em> remain lacking. In this study, we examined the effects of pH and incubation duration on lugdunin activity and evaluated the antibacterial spectrum of lugdunin produced by <em>S. lugdunensis</em> isolates against a panel of gram-positive and gram-negative bacterial strains. The optimal conditions for lugdunin antibacterial activity of isolate CGMH-SL85 were identified as pH 7.5 and a 72-h incubation period. Under the tested conditions, the lugdunin produced by CGMH-SL85 exhibited antimicrobial activity against five gram-positive strains, including <em>Staphylococcus aureus</em> ATCC29213 and <em>Staphylococcus haemolyticus</em> CGMH-SH53, followed by <em>Enterococcus faecium</em> EF029 and EF081–2 and <em>Listeria monocytogenes</em> ATCC10403S. However, no antibacterial activity was observed against any of the 11 tested gram-negative bacterial species. Furthermore, four distinct lugdunin susceptibility phenotypes were observed among 47 lugdunin-nonproducing <em>S. lugdunensis</em> strains (14 sequence type (ST)4, 27 ST27, and 6 ST29 strains), including Type A characterized by large, clear inhibition zones; Type B with smaller, clear zones; Type C displaying halo-like inhibition zones; and Type D showing no detectable activity. Moreover, 20 <em>S. lugdunensis</em> strains (42.6 %) exhibited the Type C phenotype. Notably, all six ST29 strains displayed the Type C phenotype, while the Type A phenotype was observed only among ST27 strains (3 strains). In conclusion, we developed a standardized protocol for evaluating lugdunin activity, using pH 7.5 and a 72-h incubation period, and found that different <em>S. lugdunensis</em> strains exhibited distinct lugdunin susceptibility phenotypes.</div></div>","PeriodicalId":50312,"journal":{"name":"International Journal of Medical Microbiology","volume":"321 ","pages":"Article 151678"},"PeriodicalIF":3.6,"publicationDate":"2025-09-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145119466","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Enteroinvasive Escherichia coli (EIEC) is a diarrhoeagenic E. coli pathotype that shares key virulence traits with Shigella, including the invasion plasmid (pINV). In Thailand, an outbreak caused by the EIEC serotype O8:H19—the first reported in the country—occurred in 2023, affecting over 150 patients. To elucidate the emergence, clinical relevance, and epidemiological distribution of EIEC in Thailand, we conducted a comprehensive investigation. We isolated and genomically characterised 63 isolates, comprising 28 EIEC (eight serotypes, including O96:H19 from a 2024 outbreak) and 35 Shigella (25 S. sonnei and 10 S. flexneri), along with 85 global reference strains. Comparative genomics revealed that the 2023 and 2024 EIEC outbreak isolates, along with a novel OX18:H25 EIEC lineage, harboured highly similar pINV plasmids with conserved invasion genes and complete conjugation elements. These isolates retained several biochemical traits that were more typical of commensal E. coli than classical EIEC. Limited chromosomal genome reduction—a hallmark of Shigella— was observed, which suggests that these lineages are in an early stage of adaptation toward a pathogenic lifestyle. Phylogenomic analysis showed that OX18:H25 is closely related to livestock-associated E. coli, supporting the hypothesis that pINV was recently acquired via horizontal gene transfer. These findings highlight the active circulation of putatively conjugative virulence plasmids among E. coli populations and the ongoing emergence of novel EIEC clones with epidemic-inducing potential.
{"title":"Genomic analyses of enteroinvasive Escherichia coli revealed the circulation of conjugative virulence plasmids and emergence of novel clones","authors":"Kazuhisa Okada , Warawan Wongboot , Amonrattana Roobthaisong , Nonzee Hanchanachai , Pawinee Doung-ngern , Pilailuk Akkapaiboon Okada , Thanee Wongchai , Witaya Swaddiwudhipong , Tetsuya Iida , Shigeyuki Hamada","doi":"10.1016/j.ijmm.2025.151677","DOIUrl":"10.1016/j.ijmm.2025.151677","url":null,"abstract":"<div><div>Enteroinvasive <em>Escherichia coli</em> (EIEC) is a diarrhoeagenic <em>E. coli</em> pathotype that shares key virulence traits with <em>Shigella</em>, including the invasion plasmid (pINV). In Thailand, an outbreak caused by the EIEC serotype O8:H19—the first reported in the country—occurred in 2023, affecting over 150 patients. To elucidate the emergence, clinical relevance, and epidemiological distribution of EIEC in Thailand, we conducted a comprehensive investigation. We isolated and genomically characterised 63 isolates, comprising 28 EIEC (eight serotypes, including O96:H19 from a 2024 outbreak) and 35 <em>Shigella</em> (25 <em>S. sonnei</em> and 10 <em>S. flexneri</em>), along with 85 global reference strains. Comparative genomics revealed that the 2023 and 2024 EIEC outbreak isolates, along with a novel OX18:H25 EIEC lineage, harboured highly similar pINV plasmids with conserved invasion genes and complete conjugation elements. These isolates retained several biochemical traits that were more typical of commensal <em>E. coli</em> than classical EIEC. Limited chromosomal genome reduction—a hallmark of <em>Shigella</em>— was observed, which suggests that these lineages are in an early stage of adaptation toward a pathogenic lifestyle. Phylogenomic analysis showed that OX18:H25 is closely related to livestock-associated <em>E. coli</em>, supporting the hypothesis that pINV was recently acquired via horizontal gene transfer. These findings highlight the active circulation of putatively conjugative virulence plasmids among <em>E. coli</em> populations and the ongoing emergence of novel EIEC clones with epidemic-inducing potential.</div></div>","PeriodicalId":50312,"journal":{"name":"International Journal of Medical Microbiology","volume":"321 ","pages":"Article 151677"},"PeriodicalIF":3.6,"publicationDate":"2025-09-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145158885","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-09-16DOI: 10.1016/j.ijmm.2025.151674
Zia Ud Din , Farman Ullah , Anwar Sheed Khan , Sajjad Ahmad , Azra , Aiman Waheed , Noor Muhmmad , Fawad Ali , Farhad Ali Khattak , Gulab Fatima Rani , Otavio Cabral-Marques , Ihtisham Ul Haq , Muhammad Riaz , Jody E. Phelan , Susana Campino , Taj Ali Khan , Taane G. Clark
Background
Tuberculosis (TB), caused by bacteria of the Mycobacterium tuberculosis complex (MTBC), remains a global health challenge, exacerbated by multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains.
Objectives
This study employs whole-genome sequencing (WGS) to characterise genetic mutations associated with pyrazinamide (PZA) and fluoroquinolone (FQ) resistance in MDR-TB isolates from KPK.
Methodology
MDR and pre-XDR TB samples were collected and processed at the Provincial Tuberculosis Reference Laboratory under Biosafety Level III conditions. Samples underwent microscopy, GeneXpert MTB/RIF assay, culture, and drug susceptibility testing. DNA was extracted from positive cultures and subjected to WGS. Bioinformatics tools were used to analyse sequencing data, identify resistance-associated mutations, and assess genetic diversity among isolates.
Results
Out of the 78 MTBC isolates analysed, 67 (85.9 %) were identified as MDR-TB, with 48 categorized as pre-XDR, while 11 were drug-susceptible. The isolates predominantly came from young patients (mean age: 29.5 years, SD ±12.64), with a higher proportion of female patients (61.53 %). Mutations in the pncA gene, associated with PZA resistance, were identified in 51 isolates. Resistance to fluoroquinolones was linked to mutations in the gyrA and gyrB genes in 48 isolates. WGS confirmed PZA resistance in 51 isolates, 39 (76.47 %) of which also exhibited FQ resistance.
Conclusion
Phylogenetic analysis revealed that Lineage 3 (L3) was predominant (58.97 %), followed by L4, L2, and L1 strains. The clustering of drug-resistant strains within L3 suggests ongoing localized transmission. These findings underscore the urgent need for targeted interventions, including enhanced molecular surveillance and tailored treatment strategies, to combat MDR-TB in KPK.
{"title":"Genomic insights into pyrazinamide and fluoroquinolone resistance in multidrug-resistant tuberculosis in Khyber Pakhtunkhwa, Pakistan","authors":"Zia Ud Din , Farman Ullah , Anwar Sheed Khan , Sajjad Ahmad , Azra , Aiman Waheed , Noor Muhmmad , Fawad Ali , Farhad Ali Khattak , Gulab Fatima Rani , Otavio Cabral-Marques , Ihtisham Ul Haq , Muhammad Riaz , Jody E. Phelan , Susana Campino , Taj Ali Khan , Taane G. Clark","doi":"10.1016/j.ijmm.2025.151674","DOIUrl":"10.1016/j.ijmm.2025.151674","url":null,"abstract":"<div><h3>Background</h3><div>Tuberculosis (TB), caused by bacteria of the <em>Mycobacterium tuberculosis</em> complex (MTBC), remains a global health challenge, exacerbated by multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains.</div></div><div><h3>Objectives</h3><div>This study employs whole-genome sequencing (WGS) to characterise genetic mutations associated with pyrazinamide (PZA) and fluoroquinolone (FQ) resistance in MDR-TB isolates from KPK.</div></div><div><h3>Methodology</h3><div>MDR and pre-XDR TB samples were collected and processed at the Provincial Tuberculosis Reference Laboratory under Biosafety Level III conditions. Samples underwent microscopy, GeneXpert MTB/RIF assay, culture, and drug susceptibility testing. DNA was extracted from positive cultures and subjected to WGS. Bioinformatics tools were used to analyse sequencing data, identify resistance-associated mutations, and assess genetic diversity among isolates.</div></div><div><h3>Results</h3><div>Out of the 78 MTBC isolates analysed, 67 (85.9 %) were identified as MDR-TB, with 48 categorized as pre-XDR, while 11 were drug-susceptible. The isolates predominantly came from young patients (mean age: 29.5 years, SD ±12.64), with a higher proportion of female patients (61.53 %). Mutations in the <em>pncA</em> gene, associated with PZA resistance, were identified in 51 isolates. Resistance to fluoroquinolones was linked to mutations in the <em>gyrA</em> and <em>gyrB</em> genes in 48 isolates. WGS confirmed PZA resistance in 51 isolates, 39 (76.47 %) of which also exhibited FQ resistance.</div></div><div><h3>Conclusion</h3><div>Phylogenetic analysis revealed that Lineage 3 (L3) was predominant (58.97 %), followed by L4, L2, and L1 strains. The clustering of drug-resistant strains within L3 suggests ongoing localized transmission. These findings underscore the urgent need for targeted interventions, including enhanced molecular surveillance and tailored treatment strategies, to combat MDR-TB in KPK.</div></div>","PeriodicalId":50312,"journal":{"name":"International Journal of Medical Microbiology","volume":"321 ","pages":"Article 151674"},"PeriodicalIF":3.6,"publicationDate":"2025-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145151932","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-09-06DOI: 10.1016/j.ijmm.2025.151672
Ebrahim Golmakani , Roya Sadidi , Seyed Ahmad Hashemi , Ali Haghbin , Amir Azimian
Introduction
Methicillin-resistant Staphylococcus aureus (MRSA) remains a global health threat, with livestock-associated (LA-)MRSA ST398/t011 emerging in regions with human-animal contact. This study reports the first detection of ST398/t011 in clinical isolates from Bojnurd, northeastern Iran, alongside the hospital-associated ST239/t037 clone.
Methods
From January to April 2025, 242 clinical samples from Imam Reza Hospital were screened for S. aureus. MRSA isolates underwent antimicrobial susceptibility testing (CLSI M100-2023) and molecular characterization (mecA, mecC, vanA, cfr, PVL, tsst, sec, hla; SCCmec, agr, spa typing, MLST).
Results
Of 32 positive cultures, 7 MRSA isolates were identified (ST398/t011 [n = 4]; ST239/t037 [n = 3]). All harbored mecA and exhibited resistance to β-lactams, clindamycin, and gentamicin but remained susceptible to vancomycin/linezolid. The PVL gene was detected in one ST398/t011 isolate. ST398/t011 carried SCCmec V and agr I, while ST239/t037 had SCCmec III. Isolates were recovered from wound (n = 3), blood (n = 2), eye (n = 1), and urine (n = 1) samples, with patients aged 6–48 years.
Discussion
This first report of LA-MRSA ST398/t011 in northeastern Iran highlights potential zoonotic transmission risks in this agricultural region. Co-detection of PVL in ST398/t011 and multidrug-resistant ST239/t037 underscores the need for enhanced surveillance. Limitations include the small sample size, but findings warrant further One Health investigations to assess reservoirs and transmission dynamics.
{"title":"Multiple isolation of ST398/t011 MRSA from patients in the first half of 2025 in Imam Reza Hospital, Bojnurd, North Eastern Iran","authors":"Ebrahim Golmakani , Roya Sadidi , Seyed Ahmad Hashemi , Ali Haghbin , Amir Azimian","doi":"10.1016/j.ijmm.2025.151672","DOIUrl":"10.1016/j.ijmm.2025.151672","url":null,"abstract":"<div><h3>Introduction</h3><div>Methicillin-resistant <em>Staphylococcus aureus</em> (MRSA) remains a global health threat, with livestock-associated (LA-)MRSA ST398/t011 emerging in regions with human-animal contact. This study reports the first detection of ST398/t011 in clinical isolates from Bojnurd, northeastern Iran, alongside the hospital-associated ST239/t037 clone.</div></div><div><h3>Methods</h3><div>From January to April 2025, 242 clinical samples from Imam Reza Hospital were screened for <em>S. aureus</em>. MRSA isolates underwent antimicrobial susceptibility testing (CLSI M100-2023) and molecular characterization (<em>mecA</em>, <em>mecC</em>, <em>vanA</em>, <em>cfr</em>, <em>PVL</em>, <em>tsst</em>, <em>sec</em>, <em>hla</em>; SCC<em>mec</em>, <em>agr</em>, <em>spa</em> typing, MLST).</div></div><div><h3>Results</h3><div>Of 32 positive cultures, 7 MRSA isolates were identified (ST398/t011 [n = 4]; ST239/t037 [n = 3]). All harbored <em>mecA</em> and exhibited resistance to β-lactams, clindamycin, and gentamicin but remained susceptible to vancomycin/linezolid. The <em>PVL</em> gene was detected in one ST398/t011 isolate. ST398/t011 carried SCC<em>mec</em> V and <em>agr</em> I, while ST239/t037 had SCC<em>mec</em> III. Isolates were recovered from wound (n = 3), blood (n = 2), eye (n = 1), and urine (n = 1) samples, with patients aged 6–48 years.</div></div><div><h3>Discussion</h3><div>This first report of LA-MRSA ST398/t011 in northeastern Iran highlights potential zoonotic transmission risks in this agricultural region. Co-detection of <em>PVL</em> in ST398/t011 and multidrug-resistant ST239/t037 underscores the need for enhanced surveillance. Limitations include the small sample size, but findings warrant further One Health investigations to assess reservoirs and transmission dynamics.</div></div>","PeriodicalId":50312,"journal":{"name":"International Journal of Medical Microbiology","volume":"321 ","pages":"Article 151672"},"PeriodicalIF":3.6,"publicationDate":"2025-09-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145027810","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-09-01DOI: 10.1016/j.ijmm.2025.151671
Jinming Liu , Mengyu Liu , Xiaoyan Yu , Pei-Hui Wang , Xue Guan , Wenbo Huo , Jing Zhang , Minna Cui , Xinhua Li , Xinhe Zhou , Siyu Liu , Cong Wang , Changrui Huang , Jinghua Yu
The emergence of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) as the causative agent of COVID-19 precipitated a global health crisis of unprecedented scale. SARS-CoV-2 has been shown to interfere specifically with S phase progression during early stages of infection. Nucleocapsid (N) is an important structural protein. The abundance and early presence of N suggest that the N protein may play a pivotal role in determining the fate of host cells post-infection, including in cell cycle regulation. Our observations reveal that the SARS-CoV-2 N protein actually induces S phase arrest by promoting S phase entry and simultaneously blocking exit from this phase, which is different from previous report G1/S blockage, others describe G1 and G2/M arrest. Prolonged cell cycle arrest is frequently linked to cell death, while our data suggests the N protein curtails cell proliferation, slowing down cell growth without actively triggering cell death. Intriguingly, removing the N-arm, SR-rich region, CTD, or C-tail each abolishes the N protein’s ability to suppress cell growth, whereas deletion of the NTD does not impact this capability, nor does it affect S phase arrest. All told, the SARS-CoV-2 N protein emerges as a multifunctional actor, not only driving key aspects of viral replication but also exerting significant effects on host cell physiology by modulating cell cycle progression and growth.
{"title":"SARS-CoV-2 nucleocapsid protein delays cell cycle in S-phase","authors":"Jinming Liu , Mengyu Liu , Xiaoyan Yu , Pei-Hui Wang , Xue Guan , Wenbo Huo , Jing Zhang , Minna Cui , Xinhua Li , Xinhe Zhou , Siyu Liu , Cong Wang , Changrui Huang , Jinghua Yu","doi":"10.1016/j.ijmm.2025.151671","DOIUrl":"10.1016/j.ijmm.2025.151671","url":null,"abstract":"<div><div>The emergence of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) as the causative agent of COVID-19 precipitated a global health crisis of unprecedented scale. SARS-CoV-2 has been shown to interfere specifically with S phase progression during early stages of infection. Nucleocapsid (N) is an important structural protein. The abundance and early presence of N suggest that the N protein may play a pivotal role in determining the fate of host cells post-infection, including in cell cycle regulation. Our observations reveal that the SARS-CoV-2 N protein actually induces S phase arrest by promoting S phase entry and simultaneously blocking exit from this phase, which is different from previous report G1/S blockage, others describe G1 and G2/M arrest. Prolonged cell cycle arrest is frequently linked to cell death, while our data suggests the N protein curtails cell proliferation, slowing down cell growth without actively triggering cell death. Intriguingly, removing the N-arm, SR-rich region, CTD, or C-tail each abolishes the N protein’s ability to suppress cell growth, whereas deletion of the NTD does not impact this capability, nor does it affect S phase arrest. All told, the SARS-CoV-2 N protein emerges as a multifunctional actor, not only driving key aspects of viral replication but also exerting significant effects on host cell physiology by modulating cell cycle progression and growth.</div></div>","PeriodicalId":50312,"journal":{"name":"International Journal of Medical Microbiology","volume":"320 ","pages":"Article 151671"},"PeriodicalIF":3.6,"publicationDate":"2025-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144996347","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Elizabethkingia anophelis is an emerging multidrug-resistant Gram-negative pathogen that can cause severe nosocomial infections. Although multidrug resistance complicates the clinical management of E. anophelis, the ecological impact of stress responses, including antibiotic pressure, is unclear. We demonstrated that exposure to sub-inhibitory concentrations of imipenem promoted the secretion of antibiotic-induced outer membrane vesicles (iOMVs) by E. anophelis. This study analyzed the physical and functional characteristics of iOMVs produced by a drug-resistant clinical isolate of E. anophelis treated with imipenem and assessed the potential of E. anophelis iOMVs to modulate biofilm formation in other clinically relevant Gram-negative bacteria. High-resolution imaging and biofilm assays showed that iOMVs inhibited biofilm formation and reduced biofilm density. The inhibitory effect did not affect other nosocomial pathogens such as Pseudomonas aeruginosa, Enterobacter cloacae, or Klebsiella pneumoniae. Imipenem-induced vesiculation may inadvertently impair E. anophelis’ biofilm resilience while altering microbial competition, reshaping survival dynamics in polymicrobial environments. These results demonstrate the paradoxical effect of antibiotic stress and suggest that vesicle-mediated interactions strongly affect nosocomial pathogen ecology.
{"title":"Imipenem-induced outer membrane vesicles from Elizabethkingia anophelis inhibit biofilm formation and shift nosocomial pathogen dynamics","authors":"Yuan-Ming Tsai , Ching-Ming Liu , Hsiao-Chun Chen , Tsung-Hsuan Yang , Pang-Shuo Huang , Yu-Lin Hsu , Manoj Baranwal , Ming-Hsien Chiang","doi":"10.1016/j.ijmm.2025.151670","DOIUrl":"10.1016/j.ijmm.2025.151670","url":null,"abstract":"<div><div><em>Elizabethkingia anophelis</em> is an emerging multidrug-resistant Gram-negative pathogen that can cause severe nosocomial infections. Although multidrug resistance complicates the clinical management of <em>E. anophelis</em>, the ecological impact of stress responses, including antibiotic pressure, is unclear. We demonstrated that exposure to sub-inhibitory concentrations of imipenem promoted the secretion of antibiotic-induced outer membrane vesicles (iOMVs) by <em>E. anophelis</em>. This study analyzed the physical and functional characteristics of iOMVs produced by a drug-resistant clinical isolate of <em>E. anophelis</em> treated with imipenem and assessed the potential of <em>E. anophelis</em> iOMVs to modulate biofilm formation in other clinically relevant Gram-negative bacteria. High-resolution imaging and biofilm assays showed that iOMVs inhibited biofilm formation and reduced biofilm density. The inhibitory effect did not affect other nosocomial pathogens such as <em>Pseudomonas aeruginosa</em>, <em>Enterobacter cloacae</em>, or <em>Klebsiella pneumoniae</em>. Imipenem-induced vesiculation may inadvertently impair <em>E. anophelis</em>’ biofilm resilience while altering microbial competition, reshaping survival dynamics in polymicrobial environments. These results demonstrate the paradoxical effect of antibiotic stress and suggest that vesicle-mediated interactions strongly affect nosocomial pathogen ecology.</div></div>","PeriodicalId":50312,"journal":{"name":"International Journal of Medical Microbiology","volume":"320 ","pages":"Article 151670"},"PeriodicalIF":3.6,"publicationDate":"2025-08-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144907269","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-08-08DOI: 10.1016/j.ijmm.2025.151669
Francesca McDonagh , Kate Ryan , Aneta Kovářová , Anna Tumeo , Christina Clarke , Martin Cormican , Georgios Miliotis
Objective
This study aimed to investigate the identification of blaCARBA-positive multidrug-resistant Mixta calida isolates from human hosts and to elucidate their genomic determinants in a species-wide context.
Methods
Two carbapenemase-producing M. calida isolates were received by the Galway Reference Laboratory Service in Ireland between June and July 2024. One isolate originated from a sputum sample, while the other was recovered from a routine screening rectal swab. Initial identification was performed using MALDI-ToF mass spectrometry, with genomic confirmation via 16S rRNA sequencing, digital DNA-DNA hybridization, and Average Nucleotide Identity analysis. Antimicrobial susceptibility testing was conducted using a MicroScan panel, following EUCAST and CLSI guidelines. Whole-genome sequencing, plasmid replicon typing, and antibiotic-resistance-gene and virulence-factor profiling were employed. Comparative analysis included all additional canonical M. calida genomes from NCBI database.
Results
Both Irish isolates were taxonomically placed as M. calida and exhibited multidrug resistance against penicillins, cephalosporins, monobactams and ertapenem. The acquired genes blaKPC-3, blaOXA-9, and blaTEM-122 were detected on plasmid-borne contigs, indicating horizontal acquisition. Seven plasmid replicon types were shared between the two isolates. Both plasmid replicons and acquired antimicrobial-resistance-genes (ARGs) were seldomly identified across the species. Phylogenetic inference based on core genome analysis identified a monophyletic cluster, suggesting a single introductory event.
Conclusion
This study documents a dual occurrence of blaCARBA-positive M. calida in human colonisation and infection. The findings highlight the potential for horizontal-gene-transfer to drive the emergence of multidrug-resistant profiles in the species, underscoring the need for enhanced surveillance, diagnostic precision, and targeted infection control strategies to mitigate public health risks.
Impact statement
This study reports blaESBL and blaCARBA-positive multi-drug resistant Mixta calida isolates from distinct human hosts. Genomic analysis revealed the co-occurrence of plasmid-borne resistance genes blaKPC-3, blaOXA-9, and blaTEM-122. Species-wide phylogenetic analysis grouped the two isolates into a monophyletic cluster, suggesting a single introductory event.
{"title":"Identification of blaESBL- and blaCARBA- Positive Multi-Drug Resistant Mixta calida Isolates from Distinct Human Hosts","authors":"Francesca McDonagh , Kate Ryan , Aneta Kovářová , Anna Tumeo , Christina Clarke , Martin Cormican , Georgios Miliotis","doi":"10.1016/j.ijmm.2025.151669","DOIUrl":"10.1016/j.ijmm.2025.151669","url":null,"abstract":"<div><h3>Objective</h3><div>This study aimed to investigate the identification of <em>bla</em><sub>CARBA</sub>-positive multidrug-resistant <em>Mixta calida</em> isolates from human hosts and to elucidate their genomic determinants in a species-wide context.</div></div><div><h3>Methods</h3><div>Two carbapenemase-producing <em>M. calida</em> isolates were received by the Galway Reference Laboratory Service in Ireland between June and July 2024. One isolate originated from a sputum sample, while the other was recovered from a routine screening rectal swab. Initial identification was performed using MALDI-ToF mass spectrometry, with genomic confirmation via 16S rRNA sequencing, digital DNA-DNA hybridization, and Average Nucleotide Identity analysis. Antimicrobial susceptibility testing was conducted using a MicroScan panel, following EUCAST and CLSI guidelines. Whole-genome sequencing, plasmid replicon typing, and antibiotic-resistance-gene and virulence-factor profiling were employed. Comparative analysis included all additional canonical <em>M. calida</em> genomes from NCBI database.</div></div><div><h3>Results</h3><div>Both Irish isolates were taxonomically placed as <em>M. calida</em> and exhibited multidrug resistance against penicillins, cephalosporins, monobactams and ertapenem. The acquired genes <em>bla</em><sub>KPC-3</sub>, <em>bla</em><sub>OXA-9</sub>, and <em>bla</em><sub>TEM-122</sub> were detected on plasmid-borne contigs, indicating horizontal acquisition. Seven plasmid replicon types were shared between the two isolates. Both plasmid replicons and acquired antimicrobial-resistance-genes (ARGs) were seldomly identified across the species. Phylogenetic inference based on core genome analysis identified a monophyletic cluster, suggesting a single introductory event.</div></div><div><h3>Conclusion</h3><div>This study documents a dual occurrence of <em>bla</em><sub>CARBA</sub>-positive <em>M. calida</em> in human colonisation and infection. The findings highlight the potential for horizontal-gene-transfer to drive the emergence of multidrug-resistant profiles in the species, underscoring the need for enhanced surveillance, diagnostic precision, and targeted infection control strategies to mitigate public health risks.</div></div><div><h3>Impact statement</h3><div>This study reports <em>bla</em><sub>ESBL</sub> and <em>bla</em><sub>CARBA</sub>-positive multi-drug resistant <em>Mixta calida</em> isolates from distinct human hosts. Genomic analysis revealed the co-occurrence of plasmid-borne resistance genes <em>bla</em><sub>KPC-3</sub>, <em>bla</em><sub>OXA-9</sub>, and <em>bla</em><sub>TEM-122.</sub> Species-wide phylogenetic analysis grouped the two isolates into a monophyletic cluster, suggesting a single introductory event.</div></div>","PeriodicalId":50312,"journal":{"name":"International Journal of Medical Microbiology","volume":"320 ","pages":"Article 151669"},"PeriodicalIF":3.6,"publicationDate":"2025-08-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144828174","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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