Usefulness of 3’- 5’ IS6110-RFLP genotyping and spoligotyping of Mycobacterium tuberculosis isolated in a tertiary hospital: a retrospective study detecting unsuspected epidemiological events

Abstract

A drug resistance survey involving Mycobacterium tuberculosis isolated from patients of a tertiary Hospital in the Rio de Janeiro city (RJ), Brazil, between the years 1996 and 1998 revealed a high frequency of isoniazid (HR) resistance. These isolates were revisited and genotyped. Patients came from different RJ neighborhoods and municipalities, and 70% were outpatients. Applying the 3’ and 5’ IS 6110 -RFLP and the Spoligotype genotyping methods, the clonal structure of this population was investigated obtaining a snapshot of past epidemiological events. The 3’ clusters were subsequently 5’ IS 6110 -RFLP typed. Spoligotyping was analyzed in the SITVIT2 database. Epidemiological relationships were investigated. The major lineage was T (54.4%), and SIT 53/T1 and SIT 535/T1 were the most frequent. The T1 sublineage comprises 12.8% of resistant strains and SIT 535 were assigned for 31.8% of them. Orphan patterns corresponded to 12% and 73.3% and belonged to the T lineage. One pattern was unlisted in the SITVIT2. The 5’ IS 6110 -RFLP did not confirm 3/12 of the 3’ IS 6110 -RFLP clusters. A combination of all methods decreased the number of clusters to three. Nosocomial transmission was associated with one cluster involving a hospital cupbearer. This event was suspected in a multidrug resistant-TB inpatient caregiver who harbored a mixed infection. The 3’ IS 6110 clusters were associated with HR (p=0.046). These genotypic retrospective data may reflect a fraction of more extensive recent transmission in different communities that may be corroborated by the concentration of HR patients, and may serve as a database for further evolutionary and characterization evaluation of circulating strains and together with epidemiological data favors a more effective transmission control.

Methods for Detecting Mycobacterial Mixed Strain Infections–A Systematic Review

“…In total, 26 (21.5%) of the publications reported herein included the use of IS 6110 -typing methods for detecting MSIs involving MTBC members (Chaves et al, 1999 ; Pavlic et al, 1999 ; Yeh et al, 1999 ; Braden et al, 2001 ; Du Plessis et al, 2001 ; Richardson et al, 2002 ; García De Viedma et al, 2003 , 2005 ; Allix et al, 2004 ; Das et al, 2004 ; Shamputa et al, 2004 , 2006 ; Baldeviano-Vidalón et al, 2005 ; Cox et al, 2005 , 2007 ; Van Rie et al, 2005 ; Van Der Zanden et al, 2007 ; Andrade et al, 2009 ; Mokrousov et al, 2009 ; Gardy et al, 2011 ; Navarro et al, 2011 ; Von Reyn et al, 2011 ; Adams et al, 2012 ; Cerezo et al, 2012 ; Huyen et al, 2012 ; De Almeida et al, 2019 ), 11 of which exclusively used IS 6110 -RFLP ( Supplementary Table 2 ) and one study used IS 6110 -inverse polymerase chain reaction (PCR) (Mokrousov et al, 2009 ). All the studies employed cultures in their analysis, and IS 6110 -inverse PCR was primarily used as a tool to identify strains belonging to the Beijing evolutionary lineage (Mokrousov et al, 2003 , 2009 ).…”

“…Overall, MSIs were identified in 0.4–100% of cases using IS 6110 -RFLP in these studies ( Supplementary Table 2 ), where most reports with 100% MSI detection rates involved a single patient (Pavlic et al, 1999 ; Yeh et al, 1999 ; Andrade et al, 2009 ). In addition, a recent study detected MSIs in 3 out of 17 samples from patients using two probes for IS 6110 -RFLP instead of a single probe used conventionally (De Almeida et al, 2019 ).…”

Recent progress in the genotyping of bovine tuberculosis and its rapid diagnosis via nanoparticle-based electrochemical biosensors

“…This is followed by hybridization to disclose the size of the generated fragments and the number of IS elements using chemiluminescence (originally radiolabeling). 52,53…”

Molecular epidemiology and genetic diversity of Mycobacterium tuberculosis complex in referral health centers of Bamako, Mali: What is new?

“…The T1 family and LAM-10 from lineage 4 and M. africanum 2 from lineage 6 were the most predominant strains reported in Bamako region (Togo et al, 2017). Molecular epidemiology of Mycobacterium tuberculosis complex (MTBC) was initially based on an IS6110 restriction fragment length polymorphism (RFLP) analysis (Almeida et al, 2019).…”