Search

Background: Spontaneous miscarriage, a leading health concern globally, often occurs due to various factors, including infections. Among these, Coxiella burnetii and Brucella spp. may have adverse effects on pregnancy outcomes. While previous research has established a link between infections and spontaneous miscarriage, our study aimed specifically to investigate the presence of these two pathogens in abortion samples from women who experienced spontaneous miscarriages in Iran. Our study can add to the existing knowledge by focusing on Iran, a region with a high prevalence of C. burnetii and Brucella spp. As a result, it could provide a better understanding and unique insights into the relationship of these pathogens with spontaneous miscarriages in endemic regions. Methods: From March 2021 to March 2022, a total of 728 abortion samples (including placenta and cotyledon) were collected from 409 women who had experienced spontaneous miscarriages in the provinces of Tehran, Fars, and West Azerbaijan in Iran. The specimens included 467 Formalin-Fixed Paraffin-Embedded (FFPE) and 261 fresh frozen samples. After DNA extraction from abortion samples, the quantitative real-time PCR (qPCR) assay targeted a specific fragment of the IS1111 and IS711 elements for molecular identification of C. burnetii and Brucella spp., respectively. Furthermore, the qPCR assay employing specific primers for different species was used to determine the species of Brucella. Results: Among the studied women, 1 out of 409 (0.24%) samples tested positive for Brucella spp., specifically Brucella melitensis. There were no positive specimens for C. burnetii. Conclusions: Our study contributes to understanding the potential involvement of Brucella species in spontaneous infectious abortion within endemic regions. The identification of B. melitensis in this study highlights the need for further research in this area. However, while our results suggest a relatively low or zero identification of these pathogens in our sample population, this does not rule out the possibility of undetected infections. Therefore, it is critical to acknowledge the limitations of the molecular techniques used (qPCR), which may have potential limitations such as sensitivity and specificity. Moreover, because 64.15% of our samples were FFPE, the sensitivity of the qPCR test may be reduced. These raise concerns about the accuracy of the reported prevalence rates and the potential for false positives or negatives. [ABSTRACT FROM AUTHOR]

The genetic diversity of Brucella strains has not been fully understood. To investigate this, the genetic characteristics of 64 isolates of Brucella melitensis from sheep and goats' milk were studied using random fragment length polymorphism (RFLP) and multiple locus variable-number tandem repeat analysis (MLVA-16) methods developed in Orsay, France (MLVA-16Orsay). The RFLP analysis revealed that all 64 isolates were of biovar one. The MLVA-typing showed that one sample was simultaneously infected with two strains of B. melitensis and the genotype of 65 isolate was analyzed. Four genotypes (47, 42, 43, and 63) were identified using MLVA-8 (panel 1), whereas six genotypes (138, 125, 116,108, and two unknown genotypes) were identified using MLVA11 (panels 1 and 2A). From the review of MLVA-16 (panels 1, 2A, and 2B), panel 2B showed a very high discriminatory power. Two loci of Bruc04 and Bruc30 from this panel had diversity index values higher than 0.71 and the average diversity index was 0.619. So MLVA-16Orsay 34 showed the genotype indicating a low genetic homogeneity among the isolates. The findings of MLVA genotyping of the isolates suggest that strains of B. melitensis isolated from the milk of small ruminants in Iran are most closely related to the isolates from neighboring countries of the Eastern Mediterranean group. To the best of our knowledge, this is the first study to indicate the potential use of MLVA genotyping for simultaneous detection of specimen contamination using two different B. melitensis biovars. [ABSTRACT FROM AUTHOR]

Despite the implementation of brucellosis eradication programs in Iran, this disease is still endemic and highly prevalent among ruminants in this country. The infection of small ruminants with Brucella abortus may play a significant role in the continuation of brucellosis among the herds of ruminants. This study investigated the proportion of B. abortus and Brucella melitensis in 150 samples of sheep and goat's raw milk which were obtained from Lorestan and Hamadan provinces in the western part of Iran using the PCR method. The results revealed that among the Brucella spp. positive samples, 26.5% and 73.4% of the samples were infected with B. abortus and B. melitensis, respectively. The incidence rates of B. abortus among the sheep and goats samples were 6.8% and 12.5%, respectively. There was a significant difference between goats and sheep regarding the proportion of B. abortus. Three samples (2%) (2 goats and one sheep) were simultaneously infected with both B. melitensis and B. abortus. This article renews our knowledge about the causative agent of brucellosis in small ruminants and shows that B. abortus has a relatively high prevalence among those animals in the western regions of Iran, and its role as one of the main factors of abortion among small ruminants should not be ignored. The information provided in the present study is important for the surveillance program, as eradication programs and strategies to prevent the spread of B. abortus among small ruminants that have not been vaccinated against this microorganism may be adapted accordingly. [ABSTRACT FROM AUTHOR]

Background: Brucellosis is one of the most important zoonotic diseases caused by Gram-negative bacteria belonging to the genus Brucella. Detection of Brucella species in different countries is of utmost importance. Aims: This study aimed to detect Brucella abortus and Brucella melitensis in domestic ruminant blood samples and their ticks in western Iran. Methods: Sampling was conducted on ruminants from August to September 2020 in four different counties of Kurdistan Province, including Divandareh, Marivan, Baneh, and Sanandaj. Totally, 250 blood samples were collected from 250 small ruminants. There were no ticks on the skin of six (2.4%) ruminants, and 244 ticks were isolated from 244 animals. After genomic DNA extraction from all the collected samples, quantitative polymerase chain reaction (qPCR) was performed to detect IS711 gene. Results: Based on qPCR results, Brucella genus was detected in two blood samples (0.8%) from female sheep and four ticks (1.6%) from male sheep, including three Dermacentor marginatus (1.22%) and one Rhipicephalus turanicus (0.4%). Although B. melitensis was not detected in any tick or blood sample, one tick sample (D. marginatus) was positive for B. abortus. Conclusion: Considering the positivity of ticks for brucellosis in this study, there is a possibility of Brucella transmission from infected ticks to humans and animals through tick bites, nevertheless, in order to identify the Brucella transmission relationship between ticks and animals, serological tests should be used in future studies. [ABSTRACT FROM AUTHOR]

Brucellosis is a zoonotic infectious disease in western regions of Iran, especially in Hamedan province. Following the Famenin brucellosis cohort study, the main aim of the current study was the molecular detection of Brucella species (spp.) in sheep and goats from Famenin, Hamedan, Iran. A total of 23 Brucella-seropositive samples (sheep=21 and goats=2), which had been screened from 1,660 animals in the Famenin cohort study, were used to detect Brucella-DNA using the BCSP31 target gene and IS711 locus. In total, 20 of 23 samples were positive for Brucella infection by using specific primers. Additionally, Brucella melitensis (B. melitensis) and Brucella abortus (B. abortus) were confirmed in 90% (n=18) and 10% (n=2) of positive samples, respectively. There was no sample with the co-infection of B. abortus and B. melitensis. In this study, B. abortus was isolated from one of the goat samples. This is the first report on Brucella spp. in animals in the region. It was found that B. melitensis is the dominant spp. responsible for brucellosis in animals from Famenin. Molecular techniques are reliable tools to detect Brucella infection, especially in cases without serology findings and conclusive results. [ABSTRACT FROM AUTHOR]

Background & Objective: Brucellosis is an endemic disease with a high prevalence in Iran whose highest frequency is in the western region of the country. Genetic diversity investigation is an important method to determine the epidemiological relationship of Brucella isolates in different geographical areas. Therefore, the present study aimed to investigate the genetic diversity of human Brucella melitensis (B. melitensis) strains using the Multiple Locus Variable-Number Tandem Repeat Analysis (MLVA) Typing method in the west of the country. Materials & Methods: In this study, 20 strains of isolated B. melitensis were collected from the human serum samples of suspected Brucellosis in the west of the country and were analyzed by MLVA-16 method. Results: The results showed that 3 genotype numbers 42, 43 and 47 were identified using MLVA-8 method and using MLVA-11 method genotypes 125, 138 and 111 were recognized. Also, 16 different genotypes were detected from the analysis of the isolates by MLVA-16 method which shows a high degree of polymorphism among the isolates due to the high genetic diversity of the isolates in Panel 2B loci. Conclusion: The results showed the high genetic diversity of B. melitensis isolates in the west of the country and their genetic relationship with the known strains in the neighboring countries of the Eastern Mediterranean area, as well as the importance of the MLVA method in identifying the source of infection. [ABSTRACT FROM AUTHOR]

Brucellosis is one of the most important zoonotic diseases in many regions worldwide. This study aimed to investigate the antimicrobial properties of hydro-ethanolic extracts of propolis (EEP) samples collected from six different regions of Iran against five Brucella melitensis (B. melitensis) clinical isolates causing human brucellosis and an antibiotic-resistant B. abortus vaccinal strain (RB51). Brucella clinical isolates were first carefully identified using conventional molecular typing and Brucella bio-typing methods. Different Brucella strains were then confronted with EEPs using the disk-diffusion agar method to evaluate the antimicrobial activity of each propolis extract. Chemical composition of EEPs was then determined using HPLC-DAD, and the main phenolic compounds were quantified. It was found that all EEPs displayed significant antimicrobial activities against Brucella strains, though to varying extents. All tested clinical strains were susceptible to different EEPs with inhibition zones ranging from 18 to 38 mm diameter. Interestingly, the RB51 vaccine strain was more susceptible to EEP6 (from Markazi province), compared to conventional antibiotics used in the treatment of brucellosis. Substantial differences observed in EEP antimicrobial activity could be due to their distinct botanical origins and chemical compositions as confirmed by our HPLC analysis. The promising inhibitory effect of some propolis preparations against a broad spectrum of Brucella strains points to the need for further studies in the context of systematic clinical investigations and opens up the way for the development of natural complements in support of conventional antibiotic therapy. [ABSTRACT FROM AUTHOR]

Brucellosis is a neglected zoonotic infection with a worldwide distribution and high levels of endemism in some regions, including the Middle East. In Iran, sheep and goats constitute a major part of the livestock population, often kept by small‐scale farmers for their own consumption and economic purposes. This investigation aimed at characterizing the Brucella spp. and biovars circulating in sheep and goats under smallholder farming and their potential spillover across farms. For this purpose, from two randomly selected pastoral districts of Alborz and Fars provinces in Iran, a total of 54 aborted foetuses (38 from sheep and 16 from goats) and 528 blood samples were collected from sheep (n = 435), goats (n = 77), farmers (n = 11) and dogs (n = 5). Then, serological, bacteriological and molecular characterization of Brucella isolates was performed using standard methods. Our results showed the high seroprevalence of brucellosis in pastoral districts of Fars and Alborz provinces reaching 16.3%, 11.7% and 12.7% by using the Rose Bengal plate test (RBPT), serum agglutination test (SAT) and indirect enzyme‐linked immunosorbent assay (iELISA) test, respectively. Furthermore, the results of bacterial culture, conventional biotyping and PCR analyses showed the presence of Brucella melitensis biovar 1 and 2 infections among goat, farmers and dog of the Alborz farms and B. melitensis biovars 1, 2 and 3 among sheep of the Fars farms. Among nine seropositive farmer and dog blood samples (four farmers and five dogs), only three (two farmers and one dog) were positive in both culture and PCR tests. These results stress the need to strengthen screening and control measures in small flocks of small ruminants in Iran that could be the starting point of new outbreaks at the livestock/human interface. The present study also suggests that infected dogs may further maintain the risk of exposure to Brucella pathogens in small farms and beyond. [ABSTRACT FROM AUTHOR]

Background: Brucellosis is an important zoonotic disease in Iran. There are several reservoirs for this disease in nature. There is little information in this regard in Iran. Aims: The present study investigated the prevalence and diversity of Brucella species in ovine reticuloendothelial organs including liver, spleen, intestine (mesenteric lymph nodes), and lung. This help to address the ability of the reservoir species in disease transmission to other animals through these organs. Methods: A total of 200 ovine visceral organs including 44 intestines, 51 lungs, 52 spleens, and 52 livers were collected. The presence of different Brucella species was studied using the polymerase chain reaction (PCR). Results: The results demonstrated the prevalence of Brucella species in 56 (28.4%) samples. In addition, liver and intestine were the most and the least infected organs, respectively. In general, 48 out of 56 positive samples were infected with Brucella abortus, while 10 samples were infected with Brucella melitensis. Conclusion: Eventually, the results approved the possible presence of B. abortus among domestic animals, especially sheep and cattle, and highlighted the role of sheep as reservoir hosts for its potential transfer to cattle. [ABSTRACT FROM AUTHOR]

Background: Brucellosis is a zoonotic disease and humans get the infection mostly through consumption of raw milk. Vaccination is the best way to control brucellosis and Iran uses Rev.1 vaccine in sheep and goat flocks. It is evident that the vaccine may shed through milk so it can infect humans. The objective of the present study is to assess the shedding of the vaccine in lactating ewes and its possible immunity in their lambs through milk feeding. Methods: In a two-month period post-parturition, reduced dose Rev.1 vaccine was injected to 50 parturited ewes. From the first day of vaccination, mixed milk samples were collected and continued for 2 months. Then the samples were tested by PCR method and the sera from 70 lambs of the examined ewes were tested by modified Rose Bengal test while they were feeding milk. Results: From the 6th day until the 27th day post-vaccination, PCR represented the DNA of Rev.1 in the milk samples. All the lamb’s sera were negative in the serological test. Conclusion: As the presence of Rev.1 in milk was confirmed in this study, it is important to consider the role of the vaccine strain as a risk of infection in humans. Moreover, as the serological response in the lambs was negative, it seems that the vaccine strain didn’t immunize the lambs through milk feeding so the vaccination of lambs is necessary in small ruminant’s flocks. [ABSTRACT FROM AUTHOR]

Equine brucellosis significantly impacts the health and functionality of horses, leading to complications such as bursitis infection, septic tenosynovitis, septic arthritis, and non-specific lameness resulting from joint infections. In the present study, we used the Rose Bengal plate agglutination test (RBPT), serum agglutination test (SAT), and the 2-mercaptoethanol (2-ME) assays to find equine brucellosis. From June 2018 to September 2022, 876 blood samples were randomly taken from apparently healthy racing horses in certain parts of Iran, such as Kerman, Isfahan, Tehran, Qom, and Kurdistan. DNA extraction was carried out directly on all 63 serum samples identified as seropositive through RBPT. An additional 30 seronegative serum samples were also randomly chosen for study. Bacterial culture was also done on milk, blood, and vaginal swabs taken from seropositive horses.The bacteria that were found in the samples were then put through Bruce-ladder PCR. Our results indicated that 63 (7.1%), 21 (2.3%), and 2 (0.2%) of horses were seropositive using RBPT, SAT, and 2-ME, respectively. Also, none of the 30 DNA-extracted serum samples from seronegative horses tested positive for Brucella DNA, while 44.5% (28/63) of the DNA samples from seropositive horses yielded positive results for Brucella DNA. Out of the seropositive samples, 26 had DNA from Brucella abortus and 2 had DNA from Brucella melitensis. Also, B. melitensis biovar 1 was found in two milk samples from mares in the provinces of Kerman and Isfahan. It was identified using classical biotyping, and molecular assays. It was seen that some of healthy racing horses in some parts of Iran had antibodies against Brucella. The bacteriology and PCR methodologies provide a more comprehensive and reliable means of identifying Brucella spp. infections in horse, especially when the RBPT test came back positive. This underscores the imperative for employing molecular, bacterial, and serological methods in the diagnosis and monitoring of this zoonotic infection. Additionally, this finding suggests that Brucella is being transmitted to equine hosts as a result of its presence in ruminants. The mechanism of transmission may involve interactions between infected ruminants and susceptible equines. This discovery is significant as it underscores the potential cross-species transmission of Brucella and highlights the importance of understanding and managing the spread of the pathogen in both ruminant and equine populations., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Background: Brucellosis is considered a main health concern in humans and animals. Neither familiar molecular methods nor the classical biotyping techniques are acceptable for subtyping Brucella spp. Loci containing variable number tandem repeats (VNTRs) have recently demonstrated their practicality in typing isolates from human and animal origin despite the excessive genetic homogeneity in the genus Brucella. Methods: The genotypic characteristics of sixty-six Brucella melitensis and thirty-four Brucella abortus isolates from veterinary samples and human brucellosis cases in Iran during 2014 - 2018. They were analyzed using multiple- locus variable-number tandem-repeat analysis (MLVA) which consisted of sixteen primer pairs and designed and classified as belonging to one of the three panels: panel 1 (MLVA-8: eight loci including Bruce06, Bruce08, Bruce11, Bruce12, Bruce42, Bruce43, Bruce45, and Bruce55), panel 2A (three loci including Bruce18, Bruce19, and Bruce21), and panel 2B (five loci including Bruce04, Bruce07, Bruce09, Bruce16, and Bruce30); MLVA-11 (panels 1 and 2A), and MLVA-16 (panels 1, 2A, and 2B) using BioNumerics software (Version 7.6). Results: Using panel 1, 2A, and 2B (MLVA-16), 59 genotypes with a genetic similarity coefficient ranging from 91 to 100% were obtained from the 100 Brucella spp. isolates. For all isolates, only genotype 36 and genotype 26 were obtained using panels 1 and 2A, respectively. The B. abortus isolates showed variations at 9 different genotypes, while B. melitensis isolates have been dispersed in 50 different genotypes. Bruce16 and Bruce4 showed the highest discriminatory power. Conclusions: The MLVA-16 assay appeared to be a useful and important molecular genotyping tool that is capable of proving epidemiological linkages in outbreak and trace-back investigations and is helpful in improving the effectiveness of brucellosis control programs. [ABSTRACT FROM AUTHOR]

Background: Considering the prevalence of brucellosis in Iran, it is necessary to choose a specific and sensitive laboratory method to diagnose it in a rapid and timely manner. The aim of this study was to assess the accuracy of indirect enzyme-linked immunosorbent assay (ELISA) for detecting brucellosis in humans in order to have an appropriate alternative to conventional tests such as Wright, 2ME, and commercial ELISA kits. Materials and Methods: In this study, the recombinant protein produced from the gene (omp28) bp26 Brucella melitensis was used as an antigen for coating microplate wells. A total of 124 serum samples of normal healthy individuals (n=62) and patients with acute brucellosis (n=62) approved by STA and 2 ME tests were entered into the study. The data were analyzed in SPSS (ver.18). Results: The mean age was 39.8±13.5 years in the patient group and 36.1±12.7 years in the healthy group. Furthermore, 66.1% of the patients were male and 62.9% lived in rural regions, while these figures were respectively 71% and 45.2% in the healthy group. The sensitivity of 92% and specificity of 87% and a positive predictive value of 88% and a negative predictive value of 92% and an accuracy of 90% were determined for ELISA kit used in this study. Conclusion: The ELISA diagnostic kit reacted to most of the positive human sera. However, this kit needs to be further evaluated with a larger sample size of clinical specimens from different regions and with various clinical forms of human brucellosis. [ABSTRACT FROM AUTHOR]

Brucellosis is a worldwide bacterial zoonosis caused by Brucella spp. No approved vaccine is available for human use against the disease. In this study, outer membrane vesicles (OMVs) from a Brucella melitensis biovar 1 human isolate obtained in Iran were used to immunize BALB/c mice (n = 12) by 2 intramuscular injections with a 2‐week interval. Another group of 12 mice was used as non‐vaccinated controls. Two weeks after the last vaccination, six mice of each group were sacrificed, and proliferation and interferon gamma (IFNγ) production responses of their splenocytes were evaluated following in vitro stimulation with killed Brucella cells. The other mice were challenged with the virulent B. melitensis isolate. Two weeks later, mice were killed and spleens were cultured to determine the number of the challenge strain. The results showed proliferative response and IFNγ production of splenocytes from vaccinated mice (stimulation index: 2.18 ± 0.57, and 1519.35 ± 10.70 pg/mL, respectively) were significantly higher than those of control mice (stimulation index: 1.02 ± 0.02, and 210.01 ± 17.58 pg/mL, respectively). Numbers of the challenge strain in spleens of vaccinated mice were also significantly less than those in the controls with 1.6 units of protection. Our study revealed vaccination with OMVs of the B. melitensis isolate could induce specific immune responses and protection against infection in the mouse model suggesting their potential application for active immunization against brucellosis. [ABSTRACT FROM AUTHOR]

The objective of this survey was to assess the prevalence of brucellosis with focus on Brucella abortus and Brucella melletensis in different kinds of cats in north-east of Iran. Different kinds of cats included stray, pet, and cats which were located in dairy cattle herds tested. Blood sera of 48 (34.28%) stray, 42 (30%) pet, and 50 (35.71%) cats collected from 15 industrial dairy cattle herds. They were used for rapid and tube agglutination tests such as Rose Bengal, Wright, and 2-mercaptoethanol (2-ME). The comparison among age, sex, weight, and lifestyle of cats with serological tests of brucellosis were examined statistically. Totally, eight (5.71%) cats were mild positive with low titer. Only two (1.42%) cats reacted in 2-ME. There was no statistically significant relationship between sex of cat (male vs female) and serological results (P = 0.07). However, comparison among different age groups revealed significant differences (p = 0.009). There were no significant differences between the infected cats collected from dairy cattle herds and stray cats, but the stray cats and pet cats showed the significant differences (p = 0.028). This survey showed that Brucella infection (1) mainly occurred in male and stray cats aged 1-2 years and (2) is transiently mild, and (3) negative brucellosis in pet cats represents optimal hygiene and health monitoring of these cats by the owners. So, stray cats can be a risk factor for brucellosis in human and animal populations in north-east of Iran; thus, strong precautions over stray cats are necessary. [ABSTRACT FROM AUTHOR]

Brucellosis is a zoonotic infection in livestock that induces a major public health concern in developing countries, including Iran. Despite the efforts of the Iranian veterinary organization (IVO) to control brucellosis, it is still prevalent in domestic animals. In this regard, the present study aimed to evaluate the efficiency of the control strategy used by the IVO in infected herds on serological, cultural, and molecular methods. For this purpose, blood specimens were sampled from a total of 8750 vaccinated dairy cattle in two Brucella -infected farms. These farms were recognized as positive for Brucella by a screening program. Sera were evaluated by the Rose Bengal Plate Test and Wright test analysis. Positive dairy cattle were slaughtered under IVO supervision. The remaining cattle were evaluated every 3 weeks and positive animals were slaughtered. This procedure continued until the remaining animals revealed three successive negative responses in serological tests. Several lymph nodes and milk samples were collected from 164 seropositive cattle and subjected to bacterial isolation and confirmation by Bruceladder-polymerase chain reaction. Brucella melitensis biovar 1 and RB51 vaccine strains were recovered from milk and lymph node samples, respectively. Shedding of B. melitensis in the milk of vaccinated cows is a serious problem resulting in the further spread of brucellosis. The policy of "test and slaughter" performed on infected dairy cattle farms showed their usefulness for the control of brucellosis outbreaks. For the uncontrolled spread of brucellosis in Iran, effective control of bovine brucellosis required several serological surveillances to identify infected herds, eradication of the reservoirs, and vaccination of young heifers with RB51., Competing Interests: The authors declare that they have no conflict of interest.

Brucellosis is a widespread zoonotic disease causing considerable economic and public health problems. Despite animal vaccination, brucellosis remains endemic in some areas such as Iran, especially in the western Iranian province of Hamadan. We sought to detect some of the most common virulence-associated genes in Brucella isolated from human blood cultures to determine the prevalence of some virulence genes among Brucella isolates. Fifty-seven isolates were studied from patients with a clinical diagnosis of brucellosis who referred to the Infectious Diseases Ward of Sina Hospital in Hamadan Province, Iran, between April 2013 and July 2014. Blood samples were collected for the diagnosis of brucellosis using the BACTEC blood culture system. All of these isolates were confirmed by the bcsp31 Brucella-specific gene. We detected 11 virulence-associated genes of Brucella, namely cβg, virB, znuA, ure, bvfA, omp25, omp31, wbkA, mviN, manA, and manB, which are important for the pathogenesis of this bacterium in the intracellular environment by multiplex PCR. Totally, 149 patients with a clinical diagnosis of brucellosis were enrolled in this study. Fifty-seven (38.3%) patients had positive blood cultures. On biochemical and molecular testing, all of the isolates were Brucella melitensis. Ten of the virulence genes were detected among all of the 57 isolates, but the bvf gene was detected in 53 (93%) isolates. The high prevalence of virulence-associated genes among the Brucella isolates detected in Hamadan Province, Iran, underscores the pathogenicity of this bacterium in this region. [ABSTRACT FROM AUTHOR]

Conventional and real-time PCR assays were developed for detection and identification of Brucella species in bovine and buffalo semen samples. Totally, 91 bovine and buffalo semen samples were collected from 4 major provinces of Iran in various seasons. The animals which their semen samples were collected for this study had less than 1 year old, 1 -2 years old, 2-3 years old and more than 3 years old. Samples were cultured and DNA was extracted and novel primers have been designed using the IS711 target of Brucella species for conventional PCR. Positive results of PCR have been studied for presences of Brucella abortus and Brucella melitensis. Totally, 21.56% and 14.28% of bovine and buffalo semen samples, respectively, were positive for Brucella species. Khozestan had the highest while Sistan Va Balochestan had the lowest incidences of Brucella species in studied regions. Samples which were collected in spring season had the highest rate of infection. Also, samples which were collected from less than 1 year old bovine and buffalo semen samples had the highest incidence of Brucella species. Totally, 25 (24.50%) and 4 (3.92%) bovine semen samples and 14 (15.38%) and 1 (1.09%) buffalo semen samples were positive for Brucella abortus and Brucella melitensis. [ABSTRACT FROM AUTHOR]

Brucellosis is a zoonotic disease which is characterized by reduced fertility and abortion in several species of animals, as well as humans. Camel brucellosis is caused by Brucella abortus and Brucella melitensis. To overcome the limitations posed by other techniques such as culture and serology, a sensitive technique (PCR) was employed for the detection of brucellosis in 123 camels. Findings from this PCR study indicated a total of 11.38% of blood samples as positive for Brucella spp. and 13.01% of the lymph node samples were positive for Brucella spp. In this study, 5 out of 123 (4.065%) and 3 out of 123 (2.439%) camel blood samples were positive for B. abortus and B. melitensis, respectively. Also, 4 out of 123 (3.252%) and 2 out of 123 (1.626%) camel lymph node samples were positive for B. abortus and B. melitensis, respectively. Young camels were the most commonly infected age group, while adult camels were the less often infected age group. Also, higher prevalence of brucellosis was observed in female camels. These results have indicated that PCR is a sensitive technique which could be used as a confirmatory test for the detection of brucellosis in live camels, at the same time with the lowest risk of infection of laboratory personnel. The obtained results suggest that control and eradication programs for Brucella spp. infection seem to be necessary in camels. Our findings support the power of PCR test for Brucella spp. detection in the blood and lymph node samples and it could be easily used for routine diagnosis. [ABSTRACT FROM AUTHOR]

Background: Human brucellosis is an endemic disease in many countries including Iran. Exact diagnosis of brucellosis is not just based on clinical symptoms, because it will be considered in differential diagnosis of other diseases. Therefore, defining organism in culture or identification of organism by serological and molecular methods for confirming clinical diagnosis is necessary. Our aim was to develop a diagnostic PCR assay and define the optimal clinical specimen for this test. Materials and Methods: This cross-sectional and descriptive study was from February 2011 to November 2012. Results of standard agglutination test (SAT) and specific immunoglobulin IgG and IgM by enzyme-linked immunosorbent assay (ELISA) were compared with multiplex PCR in 116 patients with suspected brucellosis referred to the Ali Ebn-e-Abitaleb hospital, Rafsanjan, Iran. Their sera were collected and tested by SAT, ELISA and multiplex PCR. DNA was extracted from serum samples and examined by multiplex PCR involving specific primers for Brucella melitensis and Brucella abortus based on IS711 in the brucella chromosome. Results: Brucellosis was confirmed in 116 patients (75% male and 25% female) based on applied diagnostic methods and clinical features. Results of ELISA, the SAT, and PCR were positive in 116, respectively. B. abortus and B. melitensis were detected in 101 and 15 patients. Conclusion: The results of present study showed that multiplex PCR assay is a rapid and sensitive technique for diagnosis of brucellosis compared to SAT. However it is more accurate when coupled with conventional methods. [ABSTRACT FROM AUTHOR]

Background: Brucellosis, a zoonosis mainly transmitted by consumption of unpasteurized dairy products as well as direct contact with infected animals, is endemic in Kyrgyzstan. However, Brucella species in humans have not been investigated and the origin of the disease remains poorly known in wide parts of Сentral Asia. Thus, molecular characterization of the circulating strains is a critical first step in understanding Brucella diversity in the country., Methods: In this study, isolates were collected from patients with suspected brucellosis from different regions in Kyrgyzstan between 2019 and 2020. The detection and identification of Brucella was carried out by Bruce-ladder PCR. Next generation sequencing was used to sequence the 89 Brucella isolates, which were genotyped by cgSNP and cgMLST to identify epidemiological connection between Brucella isolates as well as placing them in the context of the global Brucella phylogeny., Results: The Brucella strains isolated from all regions of Kyrgyzstan were identified as B. melitensis. Based on cgSNP analysis, 18 sequence types were differentiated. The highest numbers of different sequence types were found in Batken (n = 8), Osh (n = 8) and Jalal-Abad (n = 6) oblasts. According to cgSNP and cgMLST analyses, different B. melitensis lineages circulate in Kyrgyzstan, all of them belonging to the Eastern Mediterranean group of the global Brucella phylogeny with the highest similarity to strains from Turkmenistan, Iran and Turkey., Conclusion: In the present study, B. melitensis was identified as a causative agent of human brucellosis in Kyrgyzstan and different lineages could be identified. Since this study focused on isolates of human origin, the identity of Brucella species and lineages circulating among animal populations remains elusive. Implementing culture techniques and use of most recent molecular, bioinformatic and epidemiological tools are needed to set up a One Health approach to combat brucellosis in Kyrgyzstan. Further, other Сentral Asian countries need to take part in this effort as brucellosis is a transboundary disease in these regions., (© 2022. The Author(s).)

• The first comprehensive molecular typing of Brucella spp. infecting humans and livestock in Iran is presented. • Twenty –three Brucella isolates were characterized by phenotypic typing, AMOS-PCR, Bruce-ladder and MLST. • B. melitensis isolates belonged to 3 MLST-9 genotypes ST8, ST7, and ST102 – the latter previously reported only once, from a traveler with links to Iran. • Two widely dispersed MLST-9 genotypes were identified from B. abortus isolates (ST1 and ST2). • The study provides the first data describing how Brucella isolates from Iran relate to the global population structure. Brucellosis is an endemic infection in Iran and represents a serious health problem in humans and livestock causing important economic losses. The objective of this study was to undertake molecular characterization of Brucella spp. isolated from humans and livestock in several provinces of Iran including by multi-locus sequence typing (MLST), in order to understand the genotypes circulating in Iran and their relationship to genotypes globally. A total of 23 Brucella isolates were isolated from eight milk samples (seven cows, and one camel), human blood samples (seven), bovine lymph nodes (two), and samples from aborted fetuses (three sheep, two cows, and one goat). Phenotypic and molecular identification of Brucella isolates was performed on all isolated bacteria and showed that all were either Brucella melitensis or Brucella abortus. B. melitensis was associated with ovine/caprine and camel samples, most human isolates, and a significant minority of cattle isolates. In contrast B. abortus from livestock was associated only with isolations from bovine samples, as well as a single human sample. These results indicate that both B. melitensis and B. abortus contribute to the human brucellosis burden in Iran. B. melitensis isolates comprised three MLST-9 genotypes, the common and globally distributed ST8, a single representative of ST7, and several additional examples of ST102, a genotype previously only reported in a single isolate from a human brucellosis case believed to be acquired through travel to Iran. B. abortus isolates represented two globally common MLST-9 genotypes (ST1 and ST2), with relationships to biotype and other PCR-based typing methods consistent with previous observations. The results provide the basis for further studies examining the molecular epidemiology of Brucella circulating in Iran and the relationships of local isolates to those present globally. [ABSTRACT FROM AUTHOR]

Live attenuated Brucella melitensis Rev.1 strain is currently used in some countries against caprine and ovine brucellosis. Although live B. melitensis Rev.1 strain is considered the best vaccine available for the prophylaxis of brucellosis in sheep and goats, its residual abortifacient potential in pregnant animals is still a problem. We used a specific PCR-RFLP method to distinguish B. melitensis Rev.1 from other Brucella field isolates derived from aborted fetuses of sheep and goats in Iran. Molecular typing of 46 Brucella spp. isolates revealed that five strains were B. melitensis Rev.1 vaccine strain. [ABSTRACT FROM AUTHOR]

It is extremely important to investigate the presence of Brucella melitensis as a nonspecific and heterogeneous agent in dairy cows in Iran due to mixed populations of sheep, goats, and cattle. B. melitensis from infected sheep or goat herds may be introduced into the cattle population in this area. Hence, it is essential to obtain epidemiological data on the probable existence of B. melitensis as a different source of the infection or new exotic Brucella phenotypes development in dairy cow herds in Iran. The purpose of this study was to investigate the presence of B. melitensis infection in cows, using both traditional bacteriological tests and polymerase chain reaction (PCR)-based assay. Traditional biotyping and PCR results of 42 Brucella spp. isolates from Tehran and Fars (northern and southern) provinces of Iran identified five (11.9%) and 37 (88.1%) as B. melitensis (four biovar 1 and one biovar 2) and 37 (88.1%) Brucella abortus biovar 3. This study demonstrated that B. melitensis infection in dairy cows of Iran is still present at a low level. However, the Iranian Veterinary Organization should be careful to have dairy cow herds free of B. melitensis infection as a nonspecific agent of brucellosis, since this type of brucellosis may be extended and act as a potential source of severe infection in humans. [ABSTRACT FROM AUTHOR]

Sequence-based Polymerase Chain Reaction (PCR) has been introduced as an effective and reliable method for bacterial strain typing, which could provide a reliable typing approach for clinical laboratories. This study aimed to describe the reproducibility and performance of the Outer Membrane Protein 31 ( Omp31 )-based PCR, as a molecular genotyping tool for Brucella melitensis ( B. melitensis ) typing. The 31 KD outer-membrane protein of Brucella, which encodes the Omp31 gene, can be applied as an antigen to diagnose brucellosis. For this purpose, 146 samples were taken from human blood samples, bovine and camel lymph nodes, as well as sheep and goat aborted fetuses, including fetal kidney, abomasum, liver, lung, spleen, and heart for bacteriological investigation. The molecular detection of the Omp31 and IS711 genes was performed using the isolated B. melitensis (n=14). The sequencing of the Omp31 gene of B. melitensis in the Iranian field isolates was also performed for the whole gene sequencing. The homology of all sequences was then checked with the reported National Center for Biotechnology Information sequences using a basic local alignment search tool for the nucleotide diversity evaluation. The findings revealed that B. melitensis isolates were recovered from 14 examined cases and confirmed by the IS711 -based PCR with a PCR product of 731 bp. Moreover, 14 Iranian B. melitensis sequences clustered together as a monophyletic grouping with bootstrap support of 63, and they were closely related to the B. melitensis reference isolates. This Omp31 -based phylogenetic placement strongly indicates the monophyletic origin of the Iranian B. melitensis in different animals and human hosts.

Brucellosis during pregnancy is associated with major concerns for mothers but, it is still not clear whether the infection could be transmitted through breastfeeding to newborns. This study aimed at evaluating the shedding of Brucella melitensis through human milk. We describe phenotypic and genotypic characterization of Brucella isolate from human milk. The characterized isolate by Bruce-ladder PCR, AMOS PCR and biotyping confirmed the presence of Brucella melitensis biovar 1 from human milk. However, the breastfeeding of newborn baby induced no serious abnormality, although occasional weakness, loss of appetite and vomiting were reported by the parents. All Brucella serological tests including RBT, SAT and 2 ME test were also positive for the baby and her mother, although the blood culture was negative for the baby. Evaluation of the blood DNA from mother and her baby also confirm the presence of Brucella melitensis in the blood samples. Therefore, the isolation of B. melitensis biovar 1 from human milk as well as presence of Brucella melitensis in the blood samples confirms breastfeeding as a possible route for infant infection. • The first report of Brucella melitensis infection in mother and her child in Iran. • Brucella melitensis biovar 1 was confirmed from human milk by molecular and biotyping methods. • Breastfeeding as a possible route for infant infection to brucellosis was revealed. • AMOS- PCR and Bruce-ladder PCR showed that the isolate belonged to Brucella melitensis. [ABSTRACT FROM AUTHOR]

Brucellosis is a widespread zoonotic disease affecting human and livestock health. This meta-epidemiological study is aiming to draw a comprehensive picture of the Brucella prevalence in Iranian livestock, trying to estimate most affected subgroups as well as the most appropriate methods and sampling conditions for brucellosis screening programs. A literature search was performed among data published between 1 January 1970 and July 2020. Different subgroups were compared according to animal species, gender, age, sampling season, sampling locations as well as the diagnostic method used for brucellosis screening. To determine heterogeneity of studies, Chi-squared test was used and a random effect model (REM) estimated the pooled prevalence among subgroups. A total of 45 publications, comprising 240 studies/data-reports, were evaluated. A significant increase in the number of studies was found over time (Coefficient = 0.151, p value < 0.001). The most studied species in Iran was cow (n = 75), followed by sheep (n = 63), goat (n = 45), camel (n = 40) and Buffalo (n = 16). The most identified Brucella species in livestock were Brucella melitensis (n = 50), Brucella abortus (n = 39), mix infection of B. melitensis and B. abortus (n = 11) and vaccine strain of B. melitensis Rev1 (n = 4). PCR-based tests were the most common applied diagnostic method (n = 140), while the highest prevalence rate of positive samples was obtained by indirect ELISA (69%). The prevalence of brucellosis was significantly higher in females (10.91%) compared to males (8.23%). The meta-epidemiological study of brucellosis in Iranian livestock would help to strengthen surveillance, control and prevention approaches to counter the spread of this zoonotic disease. Brucella species responsible for brucellosis among livestock animals in Iran. [Display omitted] • The meta-epidemiology of brucellosis among Iranian livestock animals is presented. • Cow was the most affected livestock animal in Iran (overall prevalence of 14.7%). • The prevalence of brucellosis was higher in females (10.9%) than males (8.2%). • PCR-based methods were the main approach applied for the screening of brucellosis. • The most studied and most contaminated samples were blood and semen, respectively. [ABSTRACT FROM AUTHOR]

• The first report of Brucella melitensis biovar 2 infection in herding dog of Iran. • The results of RBT and 2ME showed a slight agreement (k = 0.159). • Brucellosis incidence was remarkably higher in dog population of Alborz province. • The role of herding dogs in the Brucella persistence among ruminant farms was revealed. Canine brucellosis is a contagious disease associated with health implications for humans as well as for a wide range of wild and domesticated animals. In this study, 173 dog blood specimens were sampled from herding dogs in three different provinces including Tehran (n = 127), Qom (n = 40) and Alborz (n = 6) provinces. The presence of Brucella antibodies was determined using Rose Bengal plate test (RBPT), slow agglutination test (SAT) and 2-mercaptoethanol (2-ME), respectively. The seropositive samples were further screened using blood culture and PCR tests to identify and differentiate the implicated Brucella species. According to our results, 24.3% (42/173), 13.8% (24/173) and 6.3% (11/173) of blood samples were tested positive using RBPT, SAT and 2-ME, respectively. However, among 42 seropositive samples, only 38.1% (16/42) and 14.2% (6/42) were positive by PCR and culture, respectively. Brucella melitensis biovar 1 and biovar 2 was isolated from the bacterial cultures of 6 blood samples and confirmed by biotyping, AMOS PCR and Bruce-ladder PCR assays. These findings highlight the potential risk of Brucella transmission from dog to humans along with other livestock and reflect the critical role of infected dogs in the persistence of Brucella infections among ruminant farms. This study stresses the need for further epidemiological investigations on canine brucellosis among herding dogs and suggests the systematic screening of the disease among companion animals such as dogs in order to improve brucellosis surveillance and control programs. [ABSTRACT FROM AUTHOR]

• The first report of Brucella melitensis isolation from seronegative camel. • Infected camel may increase the risk of brucellosis within human and livestock. • There are moderate agreements between RBT, Wright and 2ME serological results. • PCR-based methods represent a powerful approach for camel brucellosis screening. Brucellosis is a significant zoonotic infection in Iran impacting both humans and animal health status. A number of reasons such as few clinical signs complicate the diagnosis of this infection in Camelidae. Despite the ubiquitous use of serological tests for the first screening of brucellosis in camel, this approach showed several restrictions because of the intracellular properties of this organism as well as decline antibody titers in chronic stage. This study aimed at identifying the presence of Brucella spp. in blood and lymph node samples collected from slaughtered male camels of Sistan-Baluchistan province by serology, culture and polymerase chain reaction (PCR) assay. For this purpose, 2854 blood camel samples were sampled and analyzed for Brucella detection by serological screening tests. The molecular detection of IS711 gene and Bruce-ladder assay as well as culture were performed using the lymph nodes of all seropositive camel (n = 10) and 30 seronegative samples. Results showed that 0.35% (10/2854), 0.24% (7/2854) and 0.21% (6/2854) of blood samples were positive by RBPT, SAT and 2ME, respectively. However, 0% (0/10) and 70% (7/10) of lymph node samples collected from seropositive camels were positive for Brucella infection by culture and PCR, respectively. Furthermore, 6.6% (2/30) of seronegative lymph node specimens showed the presences of Brucella by PCR and culture assay. The results of the present study indicated the low seroprevalence of Brucella infection in male camels of the Sistan-Baluchistan province and highlighted the complementary role of PCR techniques for a better screening of Brucella infection among seronegative camels. Moreover, the potential shedding of Brucella within undiagnosed camel milk and secretions is a serious problem which may result in further spread and maintenance of Brucella infection among both human and livestock. Thus, for brucellosis detection and control, our results suggested that a first PCR screening supported by a bacteriological isolation on positive samples should be performed along with the serological test in endemic countries to identify the source and prevent the uncontrolled spread of the disease among camels. [ABSTRACT FROM AUTHOR]

Background: Toxin-antitoxin systems (TAs) are two-component elements, which are extensive in the bacterial genome and have a regulatory role in many cellular activities including, growth arrest, survival, biofilm formation, and bacterial persistence., Objective: TAs have not well studied in Brucella spp., Methods: We evaluated the presence of different toxin-antitoxin systems, including relE- rhhlike, Fic- Phd, Cog- Rhh, and cogT- cogAT in 40 clinical Brucella melitensis isolates using PCRbased sequencing assay., Results: Our results showed the high presence of relE-rhh-like, Fic-Phd, Cog-rhh, and cogTcogAT s TAs genes in B. melitensis isolates that were 96.25%, 92.5%, 96.25%, and 95%, respectively., Conclusion: A high presence of TAs genes in clinical B. melitensis isolates revealed that the TA system could be an antibacterial target in B. melitensis but more investigation is necessitated to elucidate the exact roles of these genes., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Background: Brucellosis is a widespread zoonotic disease with significant economic and major public health problem. Failure of response and relapse of brucellosis have reported with current therapeutic regimens. Considering the high prevalence of brucellosis and recurrence rate of this disease in Iran, widespread and inappropriate use of antibiotics could redound antibiotic resistance among Brucella isolates in the community. The aim of this study was to evaluate the antibiotic susceptibility pattern of Brucella isolates by E-test method in Hamadan, west of Iran. Methods & Materials: In this study, patients with clinical diagnosis of brucellosis who referred to Infectious Diseases Ward, Sina hospital in Hamadan were studied between April 2013 and July 2014. Blood Specimens were collected for diagnosis of brucellosis by BACTEC blood culture system. Antimicrobial susceptibility patterns of clinical isolates to gentamicin, streptomycin, rifampin, doxycycline, ciprofloxacin, ofloxacin and trimethoprimsulfamethoxazole were performed by the E-test method. Then the CLSI zone size and Minimum Inhibitory Concentration interpretive criteria were examined. Results: One hundred forty-nine patients with brucellosis were enrolled in this study. Culture of clinical samples were positive in 38.3%, of which, 91.2% were associated with positive serological test. No significant associations were found between serological tests and culture method. All clinical isolates were sensitive to doxycycline, streptomycin, gentamicin, ciprofloxacin and moxifloxacin, but intermediate sensitivity to rifampin and trimethoprim-sulfamethoxazole were found in 35.08% and 3.5% of isolates, respectively. Conclusion: Because of the high frequency of intermediate sensitivity to rifampin among brucella isolates, this drug should be prescribed with caution. We recommend restricting the use of rifampin for treatment of brucellosis except as an alternative drug for special situations. [ABSTRACT FROM AUTHOR]

The presence of Brucella melitensis and Brucella abortus genomes were investigated in the synovial fluid (SF) samples from 90 patients with rheumatoid arthritis (RA). DNA extraction and PCR assay were performed for simultaneous identification and discrimination of B. melitensis and B. abortus from the SF using three specific primers. After gel electrophoresis, the PCR products were confirmed by DNA sequencing. The cbg, omp31, manA, virB, and znuA virulence genes typing were performed by multiplex-PCR. Of the 90 samples, 14 were positive for B. melitensis (n = 9; 10%) and B. abortus (n = 5; 5.5%). The virulotyping of positive samples revealed the presence of all five virulence genes in B. melitensis. The virB, cbg, and om31 were detected in all five samples of B. abortus. In addition, zhuA and manA were detected in three (60%) and four (80%) samples, respectively, of the B. abortus-positive samples. Moreover, a total of 94.2% and 89.2% of the 14 positive samples were also found positive for manA and znuA, respectively. Our findings revealed that the Brucella spp. genomes can be detected in the SF of RA patients by the PCR-based method. We thus suggest that physicians should consider the Brucella spp. as indicators of potential RA for the timely diagnosis and treatment of RA., (Copyright © 2018. Published by Elsevier Ltd.)

Brucella is zoonotic pathogen that induces abortion and sterility in domestic mammals and chronic infections in humans called Malta fever. It is a facultative intracellular potential pathogen with high infectivity. The virulence of Brucella is dependent upon its potential virulence factors such as enzymes and cell envelope associated virulence genes. The aim of this study was to investigate the Brucella virulence factors among strains isolated from humans and animals in different parts of Iran. Seventy eight strains of Brucella species isolated from suspected human and animal cases from several provinces of Iran during 2015-2016 and identified by phenotypic and molecular methods. The multiplex-PCR (M-PCR) assay was performed in order to detect the ure, wbkA, omp19, mviN, manA and perA genes by using gene specific primers. Out of 78 isolates of Brucella spp., 57 (73%) and 21 (27%) isolates were detected as B. melitensis and B. abortus, respectively, by molecular method. The relative frequency of virulence genes ure, wbkA, omp19, mviN, manA and perA were 74.4%, 89.7%, 93.6%, 94.9%, 100% and 92.3%, respectively. Our results indicate that the most of Brucella strains isolated from this region possess high percent of virulence factor genes (ure, wbkA, omp19, mviN, manA and perA) in their genome. So, each step of infection can be mediated by a number of virulence factors and each strain may have a unique combination of these factors that affected the rate of bacterial pathogenesis., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Molecular prevalence of nine putative virulence factors in two more prevalent Brucella species in Iranian patients and livestock was investigated. During five years (2010-2015), 120 human and animal specimens were collected from three geographical areas of Iran. All samples were cultured in blood culture media and subcultured into Brucella agar medium. Nine primer pairs were designed for detection of VirB2, VirB5, VceC, BtpA, BtpB, PrpA, BetB, BPE275 and BSPB virulence factors using PCR and sequence analysis. Totally, 68 Brucella isolates including 60 B. melitensis and 8 B. abortus were isolated from the human and animal specimens examined. Approximately, all B. melitensis and B. abortus strains were positive (100%) regarding btpA, btpB, virB5, vceC, bpe275, bspB, and virB2 genes except for prpA and betB that were detected in 86% and 97% of the strains, respectively. Significant relationships were found between the presence of prpA and human B. melitensis isolates (P = 0.04), and also between the presence of betB and human isolates of B. abortus (P = 0.03). In conclusion, our results revealed that Iranian Brucella strains, regardless of human or animal sources, are extremely virulent due to high prevalence of virulence attributes in almost all strains studied., (Copyright © 2017 Elsevier Ltd. All rights reserved.)