Persister cells formation and expression of type II Toxin-Antitoxin system genes in Brucella melitensis (16M) and Brucella abortus (B19)

doi:10.30699/ijp.2020.118902.2294

. 2020 Spring;15(2):127-133.

doi: 10.30699/ijp.2020.118902.2294. Epub 2020 Feb 19.

Persister cells formation and expression of type II Toxin-Antitoxin system genes in Brucella melitensis (16M) and Brucella abortus (B19)

Fatemeh Amraei ^{1

2}, Negar Narimisa ^{1

2}, Behrooz Sadeghi Kalani ³, Vahid Lohrasbi ^{1

2}, Faramarz Masjedian Jazi ^{1

2}

Affiliations

PMID: 32215028
PMCID: PMC7081757
DOI: 10.30699/ijp.2020.118902.2294

Persister cells formation and expression of type II Toxin-Antitoxin system genes in Brucella melitensis (16M) and Brucella abortus (B19)

Fatemeh Amraei et al. Iran J Pathol. 2020 Spring.

. 2020 Spring;15(2):127-133.

doi: 10.30699/ijp.2020.118902.2294. Epub 2020 Feb 19.

Authors

Fatemeh Amraei ^{1

2}, Negar Narimisa ^{1

2}, Behrooz Sadeghi Kalani ³, Vahid Lohrasbi ^{1

2}, Faramarz Masjedian Jazi ^{1

2}

Affiliations

¹ Microbial Biotechnology Research Center, Iran University of Medical Science, Tehran, Iran.
² Department of Microbiology, School of Medicine, Iran University of Medical Sciences, Tehran, Iran.
³ Clinical Microbiology Research Center, Ilam University of Medical Sciences, Ilam, Iran.

PMID: 32215028
PMCID: PMC7081757
DOI: 10.30699/ijp.2020.118902.2294

Abstract

Background & objective: Persister cells are defined as a subpopulation of bacteria that are capable of reducing their metabolism and switching to dormancy in stress conditions. Persister cells formation has been attributed to numerous mechanisms, including stringent response and Toxin-Antitoxin (TA) systems. This study aimed to investigate the hypothetical role of TA systems in persister cells formation of Brucella strains by evaluating toxins of type II TA systems (RelE, Fic, Brn T, cogT) expression.

Methods: Brucella strains treated with a lethal dose of gentamicin and ampicillin and to determine the number of surviving cells, bacterial colonies were counted at different time intervals. The role of TA systems in persister cell formation was then determined by toxin expression levels using qRT- PCR method.

Results: Our results showed the viability of persister cells after 7 h. The results of relative qRT- PCR showed higher levels of toxin gene expression due to stress conditions, suggesting the possible role of TA systems in persister cells formation and antibiotics tolerance.

Conclusion: The results of this study showed that considering the importance of persistence and the tolerance to antibiotics, further studies on persister cells formation and related genes such as the TA system genes in Brucella strains might help us to identify the precise mechanisms leading to persister cells formation.

Keywords: Brucella abortus; Brucella melitensis; Persister cell; Real-time PCR; TA systems.

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Figures

Fig. 1

Fig. 1

The presence of TA systems detection by PCR in Brucella. Lanes 1-6 respectively contains: 50 bp DNA ladder, 1: positive control 16S rRNA gene (134 bp), 2: negative control, 3: RelE (208 bp), 4: Fic (111bp), 5: Brn T (106 bp) ,6: CogT (122 bp).

Fig. 2

Fig. 2

Stabilization test for bacteria through colony count (CFUs). (Fig. 2 A) Experiment for persister cells heritability using gentamicin and ampicillin antibiotics against Brucella melitensis (16M) and (Fig. 2 B) Brucella abortus

Fig. 3

Fig. 3

Gene expression analysis by Real-time PCR. Relative expression is normalized for Brucella melitensis (16 M) and Brucella abortus (B19) with housekeeping 16S rRNA gene (P <0.05)

See this image and copyright information in PMC

References

1. Probert WS, Schrader KN, Khuong NY, Bystrom SL, Graves MHJJocm. Real-time multiplex PCR assay for detection of Brucella spp B abortus and B. melitensis. 2004;42(3):1290–3. - PMC - PubMed
1. Eskandari-Nasab E, Moghadampour M, Hasani SS, Hadadi-fishani M, Mirghanizadeh-Bafghi SA, Asadi-Saghandi A, et al. Relationship between gamma-interferon gene polymorphisms and susceptibility to brucellosis infection. Microbiol Immunol. 2013;57(11):785–91. - PubMed
1. López-Goñi I, García-Yoldi D, Marín CM, de Miguel MJ, Barquero-Calvo E, Guzmán-Verri C, et al. New Bruce-ladder multiplex PCR assay for the biovar typing of Brucella suis and the discrimination of Brucella suis and Brucella canis. Veterinary microbiology. 2011;154(1-2):152–5. - PubMed
1. Jazi FM, Mirnejad R, Piranfar V, Mozafari NA, Salehi TZ, Khormali M, et al. Real-time PCR and high-resolution melt analysis methods for detection of pathogenic species of Brucella. 2017;41(6):325–31.
1. Irajian GR, Jazi FM, Mirnejad R, Piranfar VJIjop. Species-specific PCR for the diagnosis and determination of antibiotic susceptibilities of Brucella strains isolated from Tehran, Iran. 2016;11(3):238. - PMC - PubMed

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[1] Probert WS, Schrader KN, Khuong NY, Bystrom SL, Graves MHJJocm. Real-time multiplex PCR assay for detection of Brucella spp B abortus and B. melitensis. 2004;42(3):1290–3. - PMC - PubMed

[2] Probert WS, Schrader KN, Khuong NY, Bystrom SL, Graves MHJJocm. Real-time multiplex PCR assay for detection of Brucella spp B abortus and B. melitensis. 2004;42(3):1290–3. - PMC - PubMed

[3] Eskandari-Nasab E, Moghadampour M, Hasani SS, Hadadi-fishani M, Mirghanizadeh-Bafghi SA, Asadi-Saghandi A, et al. Relationship between gamma-interferon gene polymorphisms and susceptibility to brucellosis infection. Microbiol Immunol. 2013;57(11):785–91. - PubMed

[4] Eskandari-Nasab E, Moghadampour M, Hasani SS, Hadadi-fishani M, Mirghanizadeh-Bafghi SA, Asadi-Saghandi A, et al. Relationship between gamma-interferon gene polymorphisms and susceptibility to brucellosis infection. Microbiol Immunol. 2013;57(11):785–91. - PubMed

[5] López-Goñi I, García-Yoldi D, Marín CM, de Miguel MJ, Barquero-Calvo E, Guzmán-Verri C, et al. New Bruce-ladder multiplex PCR assay for the biovar typing of Brucella suis and the discrimination of Brucella suis and Brucella canis. Veterinary microbiology. 2011;154(1-2):152–5. - PubMed

[6] López-Goñi I, García-Yoldi D, Marín CM, de Miguel MJ, Barquero-Calvo E, Guzmán-Verri C, et al. New Bruce-ladder multiplex PCR assay for the biovar typing of Brucella suis and the discrimination of Brucella suis and Brucella canis. Veterinary microbiology. 2011;154(1-2):152–5. - PubMed

[7] Jazi FM, Mirnejad R, Piranfar V, Mozafari NA, Salehi TZ, Khormali M, et al. Real-time PCR and high-resolution melt analysis methods for detection of pathogenic species of Brucella. 2017;41(6):325–31.

[8] Jazi FM, Mirnejad R, Piranfar V, Mozafari NA, Salehi TZ, Khormali M, et al. Real-time PCR and high-resolution melt analysis methods for detection of pathogenic species of Brucella. 2017;41(6):325–31.

[9] Irajian GR, Jazi FM, Mirnejad R, Piranfar VJIjop. Species-specific PCR for the diagnosis and determination of antibiotic susceptibilities of Brucella strains isolated from Tehran, Iran. 2016;11(3):238. - PMC - PubMed

[10] Irajian GR, Jazi FM, Mirnejad R, Piranfar VJIjop. Species-specific PCR for the diagnosis and determination of antibiotic susceptibilities of Brucella strains isolated from Tehran, Iran. 2016;11(3):238. - PMC - PubMed

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Persister cells formation and expression of type II Toxin-Antitoxin system genes in Brucella melitensis (16M) and Brucella abortus (B19)

Affiliations

Persister cells formation and expression of type II Toxin-Antitoxin system genes in Brucella melitensis (16M) and Brucella abortus (B19)

Authors

Affiliations

Abstract

Figures

References

LinkOut - more resources

Full Text Sources