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Background Brucellosis, a zoonotic disease in Türkiye, which has significant direct and indirect impacts on the healthcare system and livestock. This study, which aimed to investigate the differences among Brucella spp. isolates originating from different regions of Türkiye, for implications for public health and veterinary medicine. Method Twenty-one isolates from ruminants and two isolates from humans obtained from various regions of Türkiye were utilized in the study. The isolates were identified and biotyped using traditional microbiological procedures, and whole-genome sequencing (WGS) was performed. This was followed by single nucleotide polymorphism (SNP)--based phylogenetic analysis and WGS-based analysis of virulence and resistance genes. Additionally, phenotypic antimicrobial resistance and phage susceptibilities were determined. The obtained data were then compared for concordance, ensuring the validity and reliability of the results. Results Our study, employing culture methods, polymerase chain reaction (PCR), and WGS analyses, identified 11 Brucella melitensis (bv 3 ( n  = 9), one each bv 1 and bv 2) and 12 B. abortus (bv 3 ( n  = 11), bv 9 ( n  = 1)) isolates All B. abortus isolates were of bovine origin, while the B. melitensis isolates were from sheep ( n  = 7), goat ( n  = 1), ram ( n  = 1), and humans ( n  = 2). In the whole-genome SNP-based phylogenetic tree, all B. melitensis strains were found to be of the IIb subtype of genotype II associated with the Eastern Mediterranean lineage. Ten different genotypes were identified in the SNP analysis of the isolates, with a maximum SNP difference of 278 and a minimum SNP difference of 4 among these genotypes. According to the WGS-SNP-based phylogenetic tree of B. abortus isolates, they were grouped in clade C1. In the SNP analysis, where ten different genotypes were identified, the SNP difference among these genotypes was a maximum of 316 and a minimum of 6. In the in silico MLST analysis performed with WGS data, B. melitensis isolates were identified as ST8 and ST102 genotypes, while B. abortus isolates were identified as ST2 and ST3 genotypes. The dominant genotypes were ST8 for B. melitensis and ST2 for B. abortus , respectively. Virulence gene analysis conducted based on WGS data of the 23 B. abortus and B. melitensis isolates revealed 43 virulence gene-associated regions in all strains, irrespective of species, host, or isolation year. Although classical resistance-related genes were not detected by WGS-based antimicrobial resistance gene analysis, phenotypic resistance analysis revealed resistance to azithromycin, rifampin, and trimethoprim/sulfamethoxazole in B. abortus and B. melitensis isolates. Conclusion Both B. melitensis and B. abortus were circulating species in animals and human. The dominant genotypes were ST8 for B. melitensis and ST2 for B. abortus , respectively. All B. melitensis strains were found to be of the IIb subtype of genotype II associated with the Eastern Mediterranean lineage, while B. abortus isolates, they were grouped in clade C1. Further, a comprehensive study with a sufficient number of isolates covering all regions of Türkiye would provide more accurate information about the current epidemiological situation in the country.

Summary Brucellosis, caused by Gram-negative Brucella , spreads in human and animal populations through contact with infected animals and products. Developing a rapid and sensitive detection technology for pathogen is crucial to reduce the risk of this disease transmitting between animal populations and to humans. We produced a monoclonal antibody LPS-6B5, which shows high affinity to LPS and limited cross-reactivity with other bacteria. Based on LPS-6B5, a colloidal gold immunochromatographic assay (GICA) was developed which demonstrates high sensitivity and specificity in detecting cultured B. melitensis , B. abortus and B. suis . The Gold Immunochromatographic Assay (GICA) strips exhibited the most sensitive detection limits, with a value of 7.8125 × 10 5 CFU/mL for Brucella melitensis , surpassing the sensitivity levels observed for Brucella abortus and Brucella suis . It is also suitable for clinical and field samples, providing a cost-effective and user-friendly alternative to traditional methods.

We report possible intraoperative transmission of Brucella melitensis in Slovenia, likely caused by aerosolized particles during wound irrigation. Whole-genome multilocus sequence typing revealed that isolates from the patient and the surgeon belonged to the same transmission cluster, differing by 1 allele. Our findings raise awareness of occupational risks faced by orthopedic surgeons.

Brucella melitensis is a major livestock bacterial pathogen and zoonosis, causing disease and infection-related abortions in small ruminants and humans. A considerable burden to animal-based economies today, the presence of Brucella in Neolithic pastoral communities has been hypothesised but we lack direct genomic evidence thus far. We report a 3.45X B. melitensis genome preserved in an ~8000 year old sheep specimen from Menteşe Höyük, Northwest Türkiye, demonstrating that the pathogen had evolved and was circulating in Neolithic livestock. The genome is basal with respect to all known B. melitensis and allows the calibration of the B. melitensis speciation time from the primarily cattle-infecting B. abortus to approximately 9800 years Before Present (BP), coinciding with a period of consolidation and dispersal of livestock economies. We use the basal genome to timestamp evolutionary events in B. melitensis , including pseudogenization events linked to erythritol response, the supposed determinant of the pathogen’s placental tropism in goats and sheep. Our data suggest that the development of herd management and multi-species livestock economies in the 11 th –9 th millennium BP drove speciation and host adaptation of this zoonotic pathogen. Brucella melitensis is a zoonotic bacterial pathogen of livestock that can infect humans and causes brucellosis. Here, the authors sequence an ancient specimen of B. melitensis and show that the species emerged in the Neolithic period, around the time of development of animal management practices.

Brucella melitensis is a zoonotic pathogen that poses a worldwide public health challenge. In recent years, whole-genome sequencing has become a widely accepted molecular typing method for the genomic epidemiology of brucellosis. This study reports the genomic characteristics of 24 B. melitensis strains isolated from human infections in southern Tunisia over 35 years (1988–2022). We utilized WGS to analyze the clonal relationships of these strains, their relatedness to international sequences, their antimicrobial resistance determinants, and their virulence factors. Our findings revealed a high genetic stability over three decades. All isolates were identified as B. melitensis biovar 3 and were assigned to the same sequence type, ST11, using the MLST-9 scheme. Using the MLST-21, Tunisian sequences shared 20 out of 21 alleles and were assigned to 2 closely related STs (ST89 and ST114). Phylogenetic analysis indicated that all Tunisian sequences were grouped into a single subcluster within lineage I, the West Mediterranean clade, and were highly related to other strains from the Maghreb region (Morocco and Algeria). Antimicrobial resistance analysis revealed no classical resistance determinants. However, mprF , bep CDEFG genes, and missense mutations in rpo B, gyr A, gyr B, and par C genes were identified. Virulence analysis identified 67 genes, predominantly involved in lipopolysaccharide biosynthesis and the type IV secretion system. To our knowledge, this study represents the first genomic investigation of B. melitensi s strains circulating in Tunisia. Our findings underscore the importance of genomic surveillance in understanding the epidemiology and evolution of brucellosis in North Africa.

Globally, ruminants contribute largely to the livelihood and supply of quality food for human consumption. However, small ruminants face numerous problems, including infectious diseases, in lower- and middle-income countries (LMIC). Brucellosis is one of the important zoonotic diseases affecting the range of animals caused by Brucella species, including Brucella abortus and Brucella mellitensis . Although brucellosis caused by B. mellitensis in small ruminants has never been reported in the study areas, its zoonotic importance can never be underestimated. Therefore, this study was designed to investigate the sero-molecular prevalence of B. mellitensis in small ruminants in districts Mohmand and Charsadda of Khyber Pakhtunkhwa, Pakistan. A total of 400 blood samples were collected from sheep and goats (n = 200 from each species) and analyzed by Rose Bengal precipitation test (RBPT), the indirect enzyme-linked immunosorbent assay ( i -ELISA) and polymerase chain reaction (PCR). The findings of the study indicated 13.5% and 7% of sheep while 12.5% and 12.5% of goat’s samples by RBPT and (i-ELISA) respectively. The species-specific PCR confirmed B. abortus in 70% of sheep samples and 37.5% of goat’s samples and B. mellitensis in 25% of sheep and 62.5% of goat’s samples by targeting IS711. The findings of the study concluded that B. abortus and B. melitensis were circulating in sheep and goats with a higher prevalence in the study areas. This study detected the presence of B. mellitensis for the first time in small ruminants in the study areas.

Human brucellosis is a re-emerging disease in Sichuan Province, China. In this study, bacteriology, conventional bio-typing, multi-locus sequence typing (MLST), and multiple locus variable-number tandem repeat analysis (MLVA) were applied to preliminarily characterize the strains in terms of genetic diversity and epidemiological links. A total of 101 Brucella strains were isolated from 16 cities (autonomous prefectures) from 2014 to 2021, and all of the strains were identified as Brucella melitensis bv. 3, suggesting that surveillance should focus on ruminants. MLST analysis identified four STs, namely, ST8 ( n  = 93), ST39 ( n  = 6), ST101 ( n  = 1), and ST118 ( n  = 1). The latter were new STs, indicating that strains displayed high population diversity. Six MLVA-8, namely, 42, 43, 45, 63, 83, and 114, and eight MLVA-11, namely, 111, 115, 116, 125, 180, 291, 298, and 342, genotypes were identified, demonstrating that all of the strains were from the Eastern Mediterranean lineage, and these strains exhibited a high genotype diversity. MLVA-16 analysis revealed that there was a co-existing transmission pattern, where sporadic cases and multiple outbreak events had a common origin. The dominant STs and MLVA genotypes of strains were epidemic in Northern, China, and 36 MLVA-16 genotypes were shared among strains ( n  = 51, 50.4%, 51/101) from Sichuan and strains from 22 other provinces. The findings imply that infected animals were introduced from outside the province. The surveillance and control of the disease have become public health challenges. Animal quarantines should be strengthened to prevent the spread of B. melitensis species among adjacent regions.

Human brucellosis is a re-emerging disease in Yunnan, China, that poses serious threats to public health. However, the species/biovars, geographic distribution, and genetic diversity profile of circulating strains remain unclear. In this study, bacteriology procedures, AMOS-PCR, agglutination tests with anti-A and anti-M monospecific sera, and multiple-locus variable-number tandem-repeat analysis (MLVA) genotyping approaches were applied to investigate the genetic diversity profile of the strains. A total of 514 B. melitensis strains were isolated and identified from 2017 to 2023, of which 498 were in biovar (bv.) 3 and 16 were in bv. 1; the strains were distributed in 12 cities/prefectures and 67 counties. These data show that human B. melitensis infection has become a serious public health challenge. Moreover, 514 strains were sorted into three MLVA-11 clusters (I–III): those strains harboring 15 MLVA-11 genotypes, those strains belonging to the Eastern Mediterranean lineage, and those strains exhibiting high genetic diversity. MLVA-16 was used to divide 514 strains into eight (a–h) clades that contained 208 MLVA-16 genotypes (GTs), of which 96 shared GTs comprising 402 strains, with a cluster rate of 78.21%, suggesting that the disease was potentially dominated by a cluster epidemic; however, further genome sequence analysis is necessary to uncover the epidemic pattern of the disease. The remaining 112 GTs were unique (single strain), suggesting they were linked to sporadic cases rather than a cluster epidemic. A nationwide MLVA-16 comparison showed that 81 genotypes were shared by strains from the present study and strains from 29 other provinces, implying that strains from the nationally recognized B. melitensis genotypes were continuously spread. However, further genome-based analysis is necessary to identify the transmission chain and source of infection. Another MLVA-16 comparison with global datasets showed that the strains in this study had genotypes that were completely identical to strains from Asia (Afghanistan, Kazakhstan, Mongolia, Syria, Turkey, Saudi Arabia, Vietnam, Iraq and India) and Europe (Portugal, Spain, and Greece), indicating that these strains belonged to internationally recognized lineages. Further genome analysis is needed to identify the phylogenetic relationships of these strains. The present study describes a comprehensive scenario involving bio-typing and genetic diversity characteristics of B. melitensis strains, which can provide vital clues to tailor surveillance and control measures for human brucellosis.

Brucellosis, a zoonotic disease caused by Brucella spp. globally, is of great significance not only to livestock but also to public health. The most significant of the twelve species is Brucella melitensis . This article is devoted to the endemic region of Iran and aims to uncover the molecular epidemiology of B. melitensis . Biotyping, AMOS-PCR, Bruce-ladder PCR, and the in silico method of MLVA were employed to test 40 B. melitensis isolates from humans, cows, sheep, goats, camels, and horses which are found in thirteen Iranian provinces throughout the years 2015 to 2022. The data from the MLVA-8 analysis showed that there were seven genotypes that could be identified, and the most commonly identified genotype was genotype 63. The data from the MLVA-10 analysis showed that there were seven genotypes, with genotype 213 being the most prevalent in Iran. The data from the MLVA-11 analysis showed that there were eight genotypes, with genotype 111 being the most prevalent in Iran. The MLVA and SNP analysis results showed that the bacteria were grouped into two main groups, known as the Eastern Mediterranean and American groups. Moreover, the outcomes from these analyses have added considerably to our understanding of the genetic/historical relationships among the isolates. Our study indicates a high prevalence of B. melitensis biovar 1 in Iran, accounting for 82.5% of the isolates. The study provides insight into such matters as the complex epidemiology of B. melitensis in Iran, suggesting different ways of transmission and sources of infection. This research points out the vital significance of the continuation of the surveillance and curation of B. melitensis in the diverse species of animals and humans. The simplicity and efficiency of MLVA-based molecular epidemiology offer information on the geographic distribution and genetic diversity of B. melitensis and, therefore, help in the devising of targeted strategies for the prevention of disease in animals.

In China, brucellosis is an endemic disease and the main sources of brucellosis in animals and humans are infected sheep, cattle and swine. Brucella melitensis (biovars 1 and 3) is the predominant species, associated with sporadic cases and outbreak in humans. Isolates of B. abortus, primarily biovars 1 and 3, and B. suis biovars 1 and 3 are also associated with sporadic human brucellosis. In this study, the genetic profiles of B. melitensis and B. abortus isolates from humans and animals were analyzed and compared by multi-locus variable-number tandem-repeat analysis (MLVA). Among the B. melitensis isolates, the majority (74/82) belonged to MLVA8 genotype 42, clustering in the 'East Mediterranean' group. Two B. melitensis biovar 1 genotype 47 isolates, belonging to the 'Americas' group, were recovered; both were from the Himalayan blue sheep (Pseudois nayaur, a wild animal). The majority of B. abortus isolates (51/70) were biovar 3, genotype 36. Ten B. suis biovar 1 field isolates, including seven outbreak isolates recovered from a cattle farm in Inner Mongolia, were genetically indistinguishable from the vaccine strain S2, based on MLVA cluster analysis. MLVA analysis provided important information for epidemiological trace-back. To the best of our knowledge, this is the first report to associate Brucella cross-infection with the vaccine strain S2 based on molecular comparison of recovered isolates to the vaccine strain. MLVA typing could be an essential assay to improve brucellosis surveillance and control programs.