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Objectives To establish the epidemiological correlation between the study isolates, Indian and World isolates of Acinetobacter baumannii by performing Multilocus Sequence typing. Materials and methods A total of 181 isolates (Sputum ( n  = 116), lower respiratory tract other than sputum ( n  = 36), upper respiratory tract ( n  = 20), Environment (medical) ( n  = 4), and Blood ( n  = 5) of Acinetobacter baumannii were retrieved from our repository. DNA was isolated and Multilocus Sequence Typing was performed according to the Pasteur scheme. The amplified fragments were sequenced by outsourcing, and the locus and the sequence types were determined as given in the PUBMLST site. The clonal complexes were assigned using eBURST. Results and conclusion Of the 181 isolates, 20 were colonizers and 4 were from hospital environments. All the study isolates were multidrug-resistant (MDR) and 4 of them were extensively drug-resistant (XDR). 23 sequence types were unique and were assigned new sequence types. Among them, 2125 ( n  = 12), an SLV (Single Locus Variant) of 2, was the commonest followed by 2126 ( n  = 2) which was a DLV (Double Locus Variant) of 2 and an SLV of 2125. Others were singletons. Among the known STs, 149 ( n  = 72) was the commonest followed by ST 2 ( n  = 62) & 415 ( n  = 5), ST 10 ( n  = 4), ST 15, ST622 and ST1482 (3 each). ST149 had 1SLV ST1482 ( n  = 3). ST 2 has 5 SLVs (415, 1555, 2125, 2128, & 2131, and 2 DLVs (2130 & 2126). eBURST analysis of the study isolates showed three groups: Group I (86 isolates) with ST 2 as the primary founder, group II (6 isolates) and group III (79 isolates) with ST 149 as the primary founder. All the other 11 isolates were singletons. There was no difference in antimicrobial sensitivity or sequence types of clinical and colonizing isolates. The sequence types of study isolates were compared to the world isolates in the PUBMLST database. To conclude, MLST is an important tool for establishing isolates’ phylogenetic relationships. Acinetobacter baumannii being an important nosocomial pathogen should be routinely screened for the frequent changes in the sequence types to demonstrate the emerging resistance patterns and other changes.

Yogurt, a globally consumed fermented dairy product, is recognized for its taste and potential health benefits attributed to probiotic bacteria, particularly Streptococcus thermophilus. In this study, we employed Multilocus Sequence Typing (MLST) to investigate the genetic diversity and phylogenetic relationships of 13 S. thermophilus isolates from traditional Turkish yogurt samples. We also assessed potential correlations between genetic traits and geographic origins. The isolates were identified as S. thermophilus using VITEK® MALDI-TOF MS, ribotyping, and 16S rRNA analysis methods. MLST analysis revealed 13 different sequence types (STs), with seven new STs for Turkey. The most prevalent STs were ST/83 (n = 3), ST/135 (n = 2), and ST/134 (n = 2). eBURST analysis showed that these isolates mainly were singletons (n = 7) defined as sequence types (STs) that cannot be assigned to any group and differ at two or more alleles from every other ST in the sample. This information suggests that the isolates under study were genetically distinct from the other isolates in the dataset, highlighting their unique genetic profiles within the population. Genetic diversity analysis of ten housekeeping genes revealed polymorphism, with some genes showing higher allelic variation than others. Tajima’s D values suggested that selection pressures differed among these genes, with some being more conserved, likely due to their vital functions. Phylogenetic analysis revealed distinct genetic diversity between Turkish isolates and European and Asian counterparts. These findings demonstrate the genetic diversity of S. thermophilus isolates in Turkish yogurt and highlight their unique evolutionary patterns. This research contributes to our understanding of local microbial diversity associated with yogurt production in Turkey and holds the potential for identifyic strains with enhanced functional attributes.

is primarily transmitted via contaminated food and can cause listeriosis, an infection often associated with sepsis and meningitis in at-risk individuals. Accurate outbreak detection relies on whole genome sequencing (WGS) and core genome multilocus sequence typing (cgMLST), which use allele thresholds to identify related strains. This study investigated mutation rates in , focusing on isolates with DNA repair deficiencies. Serial subcultivations were performed, comparing a repair-deficient isolate with a wild-type control. Genetic variability was assessed using WGS and cgMLST. Mutation rates were significantly higher in repair-deficient isolates, exceeding typical cgMLST thresholds currently used in outbreak investigations, leading to a misclassification of related isolates as unrelated. An additional analysis of the Austrian database revealed that such deficiencies are rare among isolates. The standard 7-allele cgMLST threshold effectively identifies related strains in most cases, but may require adjustments for hypermutator strains. Incorporating DNA repair data could improve the accuracy of outbreak investigations, ensuring reliable public health responses.

Candida auris, Candida blankii, and Kodamaea ohmeri have been regarded as emerging fungal pathogens that can cause infections with high mortality. For genotyping of C. auris, a multilocus sequence typing (MLST) scheme based on four locus sequences has been reported, while there is no typing scheme for C. blankii and K. ohmeri. In the present study, the existing MLST scheme of C. auris was modified by adding more locus types deduced from sequence data available in the GenBank database. Furthermore, MLST schemes of C. blankii and K. ohmeri were developed using the four cognate loci (ITS, RPB1, RPB2, D1/D2) and similar sequence regions to those of C. auris. These MLST schemes were applied to identify the ST (sequence type) of clinical isolates of C. auris (n = 7), C. blankii (n = 9), and K. ohmeri (n = 6), derived from septicemia or otomycosis in Bangladesh in 2021. All the C. auris isolates were classified into a single ST (ST5) and clade I, having a Y132F substitution in ERG11p, which is associated with azole resistance. Similarly, all the C. blankii isolates belonged to a single type (ST1). In contrast, six K. ohmeri isolates were assigned to five types (ST1-ST5), suggesting its higher genetic diversity. These findings revealed the availability of MLST schemes for these three fungal species for understanding their clonal diversity among clinical isolates.

Background Campylobacter ( C. ) jejuni is the leading cause of human campylobacteriosis worldwide. We performed a molecular epidemiological study to investigate the genetic relationship among C. jejuni strains isolated from human diarrhoeal patients, broiler products and dairy cattle in Lithuania. Methods The C. jejuni isolates from human clinical cases, dairy cattle and broiler products were genotyped using multilocus sequence typing (MLST). Allele numbers for each housekeeping gene, sequence type (ST), and clonal complex (CC) were assigned by submitting the DNA sequences to the C. jejuni MLST database ( http://pubmlst.org/campylobacter ). Based on the obtained sequence data of the housekeeping genes a phylogenetic analysis of the strains was performed and a minimum spanning tree (MST) was calculated. Results Among the 262 C. jejuni strains (consisting of 43 strains isolated from dairy cattle, 102 strains isolated from broiler products and 117 clinical human C. jejuni strains), 82 different MLST sequence types and 22 clonal complexes were identified. Clonal complexes CC21 and CC353 predominated among the C. jejuni strains. On ST-level, five sequence types (ST-5, ST-21, ST-50, ST-464 and ST-6410) were dominating and these five STs accounted for 35.9% ( n  = 94) of our isolates. In addition, 51 (19.5%) C. jejuni strains representing 27 (32.9%) STs were reported for the first time in the PubMLST database ( http://pubmlst.org/campylobacter ). The highest Czekanowski index or proportional similarity index (PSI) was calculated for C. jejuni strains isolated from human campylobacteriosis cases and broiler products (PSI = 0.32) suggesting a strong link between broiler strains and human cases. The PSI of dairy cattle and human samples was lower (PSI = 0.11), suggesting a weaker link between bovine strains and human cases. The calculated Simpson’s index of all C. jejuni isolates showed a high genetic diversity (D = 0.96). Conclusion Our results suggest that broiler products are the most important source of human campylobacteriosis in Lithuania. The study provides information on MLST type distribution and genetic relatedness of C. jejuni strains from humans, broiler products and dairy cattle in Lithuania for the first time, enabling a better understanding of the transmission pathways of C. jejuni in this country.

Background Salmonella Wandsworth is a rare serotype of Salmonella. This study analyzed the genotyping, genome structure, and molecular biological functions of Salmonella Wandsworth based on the results of multilocus sequence typing and next‐generation sequencing genome assembly analysis. Methods Serological typing was performed using the slide‐agglutination method. The micro broth dilution method was used to test antibiotic susceptibility. Multilocus sequence typing (MLST) was used to perform the homology analysis, while the second‐generation sequencing genome analysis was used to analyze the whole genome of the bacteria. Results Salmonella Wandsworth is Group Q Salmonella. The MLST of this strain was ST1498. Salmonella Wandsworth was sensitive to antibiotics, such as ceftriaxone, imipenem, chloramphenicol, and colistin, but was resistant to ampicillin, cefalotin, gentamicin, and ciprofloxacin. The second‐generation sequencing results showed that the genome sequence length of the bacteria was 5109457bp. Annotated COG library analysis generated 3,746 corresponding genes. After the comparison with the KEGG library, 1,340 genes, which participate in 19 types of metabolic pathways, were obtained. A total of 249 pathogenic factors and 2 disease islands were predicted. 2 CRISPR sites and 8 Cas sites were predicted. It can be seen from the evolutionary tree that Salmonella Wandsworth MLST1498 and Paratyphi B str.SPB7 are gathered together. We identified one resistance gene, namely, aac(6’)‐Iaa accounting for aminoglycoside resistance. Conclusion Salmonella Wandsworth isolated in this study is Salmonella group Q. Consequently, it is necessary to strengthen the understanding of clinical infections of Salmonella Wandsworth and carry out continuous monitoring and research. Salmonella Wandsworth is a rare serotype of Salmonella. Based on genome sequences, We employed the alignment and assembly‐free (AAF) method to construct a genomic evolutionary tree.

Paenibacillus larvae causes the American foulbrood (AFB), a highly contagious and devastating disease of honeybees. Whole-genome sequencing (WGS) has been increasingly used in bacterial pathogen typing, but rarely applied to study the epidemiology of P. larvae . To this end, we used 125 P. larvae genomes representative of a species-wide diversity to construct a stable whole-genome multilocus sequence typing (wgMLST) scheme consisting of 5745 loci. A total of 51 P. larvae isolates originating from AFB outbreaks in Slovenia were used to assess the epidemiological applicability of the developed wgMLST scheme. In addition, wgMLST was compared with the core-genome MLST (cgMLST) and whole-genome single nucleotide polymorphism (wgSNP) analyses. All three approaches successfully identified clusters of outbreak-associated strains, which were clearly separated from the epidemiologically unlinked isolates. High levels of backward comparability of WGS-based analyses with conventional typing methods (ERIC-PCR and MLST) were revealed; however, both conventional methods lacked sufficient discriminatory power to separate the outbreak clusters. The developed wgMLST scheme provides an improved understanding of the intra- and inter-outbreak genetic diversity of P. larvae and represents an important progress in unraveling the genomic epidemiology of this important honeybee pathogen.

Background Anaplasma phagocytophilum is a Gram-negative obligate intracellular bacterium that replicates in neutrophil granulocytes. It is transmitted by ticks of the Ixodes ricinus complex and causes febrile illness in humans and animals. We used multilocus sequence typing (MLST) and ankA gene-based typing to study the molecular epidemiology of the A. phagocytophilum strains circulating in a German cattle herd over one pasture season. The aim was to investigate whether co-infection with two distinct variants, reinfection with the same and/or superinfection by a different strain occurred during one pasture season. Eight genetic loci were sequenced in 47 PCR-positive samples from 15 animals. Results Five different sequence types (ST) and four ankA alleles were detected in the cattle herd. Three different ST caused clinically overt tick-borne fever in primary infected animals. The concordance between ST and ankA allele was 100%. Therefore, the housekeeping genes used for MLST and the highly variable ankA gene were concatenated to increase resolution. Co-infection could be proven because samples of chronologically close collection dates were included. Co-infecting A. phagocytophilum strains differed by 14 to 18 single nucleotide polymorphisms (SNPs). Most superinfecting variants varied by 14 SNPs from the previous strain and appeared in median after a free interval of 31 days. Thus, it is unlikely that superinfecting strains arose by in-animal evolution. Immunity against re- or superinfection was assumed because the cattle developed clinical signs only during primary infection. Conclusions The tick-pathogen-vertebrate host interaction is probably much more complex than previously thought taking into account the frequently occurring events of co-infection, reinfection and superinfection. This complex situation could not be easily simulated in an experimental infection and underlines the value of field studies.

Microsporum canis is considered one of the most common zoophilic dermatophyte species causing infections in animals and humans worldwide. However, molecular epidemiological studies on this dermatophyte are still rare. In this study, we aimed to analyse the population structure and relationships between M. canis strains (n = 66) collected in southern Italy and those isolated from symptomatic and asymptomatic animals (cats, dogs and rabbits) and humans. For subtyping purposes, using multilocus sequence typing (MLST) and multilocus microsatellite typing (MLMT), we first used a limited set of strains to screen for variability. No intraspecies variability was detected in six out of the eight reference genes tested and only the ITS and IGS regions showed two and three sequence genotypes, respectively, resulting in five MLST genotypes. All of eight genes were, however, useful for discrimination among M. canis, M. audouinii and M. ferrugineum. In total, eighteen microsatellite genotypes (A–R) were recognized using MLMT based on six loci, allowing a subdivision of strains into two clusters based on the Bayesian iterative algorithm. Six MLMT genotypes were from multiple host species, while 12 genotypes were found only in one host. There were no statistically significant differences between clusters in terms of host spectrum and the presence or absence of lesions. Our results confirmed that the MLST approach is not useful for detailed subtyping and examining the population structure of M. canis, while microsatellite analysis is a powerful tool for conducting surveillance studies and gaining insight into the epidemiology of infections due to this pathogen.

Background: A carbapenem-resistant Enterobacteriaceae (CRE) outbreak occurred in an advanced emergency medical service center [hereafter referred to as the intensive care unit (ICU)] between 2016 and 2017. Aim: Our objective was to evaluate the infection control measures for CRE outbreaks. Methods: CRE strains were detected in 16 inpatients located at multiple sites. Environmental cultures were performed and CRE strains were detected in 3 of 38 sites tested. Pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and detection of β-lactamase genes were performed against 25 CRE strains. Findings: Molecular typing showed the PFGE patterns of two of four Klebsiella pneumoniae strains were closely related and the same MLST (ST2388), and four of five Enterobacter cloacae strains were closely related and same MLST (ST252). Twenty-three of 25 CRE strains harbored the IMP-1 β-lactamase gene and 15 of 23 CRE strains possessed IncFIIA replicon regions. Despite interventions by the infection control team, new inpatients with the CRE strain continued to appear. Therefore, the ICU was partially closed and the inpatients with CRE were isolated, and the ICU staff was divided into two groups between inpatients with CRE and non-CRE strains to avoid cross-contamination. Although the occurrence of new cases dissipated quickly after the partial closure, a few months were required to eradicate the CRE outbreak. Conclusion: Our data suggest that the various and combined measures that were used for infection control were essential in stopping this CRE outbreak. In particular, partial closure to isolate the ICU and division of the ICU staff were effective.