Transmission of drug-resistant Mycobacterium tuberculosis isolates between Finnish- and foreign-born patients, 2014-2021: a molecular epidemiological study

doi:10.21203/rs.3.rs-2955449/v1

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Research Article

Transmission of drug-resistant Mycobacterium tuberculosis isolates between Finnish- and foreign-born patients, 2014-2021: a molecular epidemiological study

https://doi.org/10.21203/rs.3.rs-2955449/v1

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Background

Data on the molecular epidemiological characteristics and transmission of drug-resistant Mycobacterium tuberculosis (MTB) isolates in low-incidence settings with immigration from high-incidence settings are limited.

Method

We included 101 drug-resistant MTB isolates with whole-genome sequencing data isolated in Finland between 2014 and 2021. Mutations associated with resistance to first- and second-line drugs were identified. Potential transmission clusters were identified using a threshold of 12 single-nucleotide polymorphisms (SNPs). Highly related clusters were identified using a threshold of 5 SNPs.

Result

Of the 101 drug-resistant MTB isolates, 29 (28.7%) isolates were from Finnish-born persons and 72 (71.3%) were from foreign-born persons. The proportion of foreign-born persons with multidrug-resistant (MDR) MTB (25/72, 34.7%) was higher than that in Finnish-born persons (6/29, 20.7%). The most common mutation associated with resistance to INH and RIF was katG Ser315Thr (67/83, 80.7%) and rpoB Ser450Leu (29/35, 82.9%), respectively. Lineage 4 (36/101, 35.6%) and lineage 2 (35/101, 34.6%) were prevalent lineages. A total of 16 (15.8%) isolates were classified into 7 potential transmission clusters (≤ 12 SNPs). Furthermore, one highly related cluster (≤ 5 SNPs) was identified, including one MDR isolate from a Finnish-born person and another MDR isolate from a person born in eastern Europe.

Conclusion

Transmission of drug-resistant MTB isolates between native and foreign-born persons is limited in Finland. Monitoring transmission dynamics of drug-resistant TB is of importance to inform tuberculosis management and control in low-incidence settings with increasing immigration.

Molecular Epidemiology

Mycobacterium tuberculosis

Drug resistance

Mutation

Whole genome sequencing

Genetic cluster

Tuberculosis (TB) remains a major global health threat. According to the World Health Organization (WHO), the TB incidence (new cases per 100 000 population per year) rate rose by 3.6% between 2020 and 2021, reversing declines of about 2% per year for most of the previous two decades¹. A further concern is that the burden of drug-resistant TB (DR-TB) increased between 2020 and 2021, especially rifampicin-resistant TB (RR-TB) / multidrug-resistant TB (MDR-TB)¹.

Despite the low TB incidence in Western Europe, native populations are at increased risk of TB due to the growing immigrant population from countries with high TB incidences². Surveillance data indicated that TB patients of foreign origin represent a large majority in some low-incidence settings: Sweden (89.1%), Norway (82.8%), the UK (75.1%)³. Furthermore, MDR-TB is more prevalent among immigrants (‘foreign-born persons’) than the native population in some low-incidence countries. Immigrants comprised 96.0% of MDR-TB cases in Norway, 93.9% in Switzerland, and 75.0% in western France^4–6. TB control efforts of low-incidence regions can be compromised due to transmission events between native patients and immigrants with drug-resistant Mycobacterium tuberculosis (MTB) isolates, especially MDR-TB isolates. Some studies have reported transmission events between native patients and foreign patients with MDR-TB isolates in Germany, Switzerland, Slovakia, and Belgium using whole-genome sequencing (WGS)^7–10. A Finnish study that combined epidemiological information with Mycobacterial Interspersed Repetitive Unit Variable Tandem Repeat (MIRU-VNTR) typing data between 2014 and 2017 revealed that TB transmission between the foreign and Finnish-born population did happen, but was uncommon¹¹. MIRU-VNTR typing may have insufficient resolution to identify the transmission of MTB in low TB incidence settings. WGS is more discriminatory than MIRU-VNTR typing to understand the molecular characteristics of MTB and to identify transmission between strains¹². Further, WGS is able to detect mutations, which are undetectable by traditional molecular methods, and has become the leading technology in resistance profiling and contact tracing¹³. However, published WGS-based data on transmission of drug-resistant MTB isolates among Finnish and foreign-born patients is limited.

In this study, we included 101 drug-resistant MTB isolates with WGS data collected from 2014 to 2021 in Finland to identify the molecular characteristics of drug-resistant MTB isolates and transmission between Finnish- and foreign-born patients.

Study population

The annual number of new TB cases in Finland decreased from 264 in 2014 to 173 in 2021, with a decreasing proportion of MDR-TB from 3.0% (8/264) to 1.7% (3/173) during the same period¹⁴. The number of foreign-born persons living in Finland increased from 321 977 in 2014 to 442 290 in 2021¹⁵. Notification of TB cases to the National Infectious Disease Register (NIDR) is mandatory for laboratories and clinicians in Finland. Clinical microbiology laboratories notify new TB cases directly to the NIDR and submit isolates to the Mycobacterial Reference Laboratory at the National Institute for Health and Welfare (THL) for drug susceptibility testing (DST) and genotyping. In addition, physicians notify clinically confirmed TB cases to the NIDR using European Centre for Disease Prevention and Control criteria. The most common countries of origin among foreign born TB cases from high incidence countries in Finland were Somalia, Afghanistan and Vietnam. It is mandatory to offer TB screening by CXR and an interview within 2 weeks of arrival to asylum seekers and refugees from high TB incidence countries (≥ 50/100 000).

Phenotypic drug-susceptibility testing

Culture-based DST was performed using the automated Mycobacteria Growth Indicator Tube (MGIT; Becton Dickinson, Franklin Lakes, MD, USA) system according to the manufacturer's instructions with drug kits for I-line (streptomycin, isoniazid, rifampicin and ethambutol and pyrazinamide) and drug powder for II- and III- line testing (amikacin, capreomycin, kanamycin, ofloxacin, ethionamide, clofazimine, linezolid and moxifloxacin), using growth from solid media.

Whole-genome sequencing

Genomic DNA of MTB isolates was extracted using Magattract DNA extraction kit (Qiagen, Hilden, Germany). A 300-bp paired-end library was constructed for each DNA sample and sequenced on an Illumina (San Diego, CA, USA) platform at THL or LGC Genomics (GmbH, Berlin, Germany) for WGS according to the manufacturer’s instructions. Raw sequencing reads were subjected to quality control with Trimmomatic.

Bioinformatic analysis

This study employed TB-profiler to annotate gene mutations associated with drug resistance based on fastq. files. The trimmed and filtered reads were mapped to the MTB reference genome (H37RV, GenBank NC_000962.2) using Bowtie2 version 2.5.0. SAMtools (version 1.12) was used to sort and merge alignments. The Genome Analysis Toolkit (GATK, version 4.2.6.1) was used for single nucleotide polymorphisms (SNPs) calling. SNPs for all MTB isolates were also used to identify genetic lineage by fast-lineage-caller (version 0.3.1) and to generate a maximum likelihood based phylogenetic tree by MEGA5. Pairwise SNPs distances were calculated and cluster analysis was performed in R (version 4.2.2). Genomic clusters were defined at the following two levels: ≤5 SNPs (highly related) and 6–12 SNPs (potentially related), as these thresholds are widely accepted and used internationally¹².

Inclusion and characteristics of samples

A total of 1816 TB cases were notified between 2014 and 2021 in Finland, and 1440 of these were culture positive. Furthermore, 146 cases were identified as DR-TB using DST. Following DST, isolates from 39 cases were missing for WGS. Among sequenced isolates from 107 cases, sequencing data of 115 isolates from 103 cases were available. Among these, two isolates from two cases were excluded due to potential laboratory contamination or mixed infection, and 12 isolates were excluded due to duplicate samples. Finally, 101 first isolates from 101 DR-TB cases were included in the analysis. Among 101 DR-TB cases, 80 (79.2%) had pulmonary TB. Of 101 MTB isolates, 29 (28.7%) were from Finnish-born persons and 72 (71.3%) were from foreign-born persons. The proportion of drug-resistant TB cases varied across study years (19.8% in 2014, 8.9% in 2015, 5.9% in 2016, 16.8% in 2017, 11.9% in 2018, 13.9% in 2019, 13.9% in 2020, and 8.9% in 2021). Overall, 42 (41.6%) isolates were phenotypically assessed as mono-resistant, 1 (0.9%) as RIF mono-resistant, 25 (24.8%) as INH mono-resistant, and 16 (15.8%) as STR mono-resistant. In addition, 59 (58.4%) isolates were resistant to at least two anti-tuberculosis agents, including 25 (24.8%) poly-drug resistant MTB isolates, 31 (30.7%) MDR isolates, and 3 (3.0%) XDR isolates. The proportion of MDR (25/72, 34.7%) in foreign-born persons with drug-resistant MTB was higher than that in Finnish-born persons (6/29, 20.7%) with drug-resistant MTB (Table 1).

Table 1

Characteristics of 101 phenotypically drug-resistant Mycobacterium tuberculosis isolates in Finland 2014–2021 by region of birth
Characteristics	No. of all isolates	Region of birth
Characteristics	No. of all isolates	Finnish-born (N = 29)	Foreign-born (N = 72)	P-value
Collection year
2014	20 (19.8)	7 (24.1)	13 (18.1)
2015	9 (8.9)	1 (3.4)	8 (11.1)
2016	6 (5.9)	1 (3.4)	5 (6.9)
2017	17 (16.8)	2 (6.9)	15 (20.8)
2018	12 (11.9)	6 (20.7)	6 (8.3)
2019	14 (13.9)	3 (10.3)	11 (15.3)
2020	14 (13.9)	5 (17.2)	10 (13.9)
2021	9 (8.9)	4 (13.8)	4 (5.6)	0.218
Phenotypic drug resistance profile
RIF mono-resistance	1 (0.9)	1 (3.4)	0
INH mono-resistance	25 (24.8)	8 (27.6)	17 (23.6)
STR Mono-resistance	16 (15.8)	8 (27.6)	8 (11.1)
Poly-resistance	25 (24.8)	4 (13.8)	21 (29.2)
MDR	31 (30.7)	6 (20.7)	25 (34.7)
XDR	3 (3.0)	2 (6.9)	1 (1.4)	0.041^*
Phenotypic resistance to specific drug
RIF	35 (34.6)	9 (31.0)	26 (36.1)
INH	84 (82.1)	20 (69.0)	64 (88.9)
EMB	11 (10.9)	1 (3.4)	10 (13.9)
PZA	31 (30.7)	7 (24.1)	24 (33.3)
STR	68 (68.3)	19 (65.5)	49 (68.1)
Aminoglycosides ^a	10 (9.9)	2 (6.9)	8 (11.1)
Fluoroquinolones ^b	3 (3.0)	0	3 (4.2)	-
Abbreviations: RIF, rifampicin; INH, isoniazid; STR, streptomycin; MDR, multidrug resistance; XDR, extensive drug resistance. P-value was calculated by χ² test All values are n (%) unless otherwise indicated. ^a Amikacin and kanamycin. ^b Levofloxacin and moxifloxacin. ^* p < 0.05.

Mutations associated with drug resistance

Among 83 MTB isolates with resistance to INH, the most prevalent mutation associated with drug-resistance was katG Ser315Thr (67/83, 80.7%). There were three INH-resistant strains with inhA Ile194Thr. The most common mutation associated with STR resistance was rpsL Lys43Arg (43/69, 62.3%). Five mutations were identified (rpoB Ser450Leu, rpoB His445Asp, rpoB Leu452Pro, rpoB Gln761Asp, rpoB Val170Phe) among the 35 isolates resistant to RIF. The most prevalent RIF-resistance associated mutation was rpoB Ser450Leu (29/35, 82.9%). Among 11 MTB strains with resistance to EMB, eight carried a mutation of embB Met306Val (8/11, 72.7%). Of 31 phenotypically PZA-resistant isolates, 80.6% (25/31) carried variation in pncA while six isolates did not. A total of 20 genetic variants associated with PZA phenotypic resistance were found. There were three isolates with FQs resistance, of which two FQs-resistant isolates carried gyrA Asp94Ala, and one carried gyrA Ala90Val (Table 2). One isolate with katG mutation (Ala110Val) was INH phenotypically susceptible. The mutation of embB Gln497Lys was found in one phenotypically EMB-susceptible isolate. Two isolates with mutation on gid were phenotypically susceptible to STR, one on Tyr22* and another on Gly173*. In addition, gyrA Asp94Gly and gyrA Asp94Ala were respectively found in two isolates with susceptibility to FQs (Table 3).

Table 2

Mutations potentially associated with drug-resistance genes in 101 phenotypically drug-resistant MTB isolates in Finland between 2014 and 2021
Isolates with phenotypic resistance to (N)	Gene	Mutation	Amino acid substitution	Frequency
INH (84)	inhA	581T-C	Ile194Thr	3
INH (84)	katG	944G-C	Ser315Thr	67
RIF (35)	rpoB	1349C-T	Ser450Leu	29
		1333C-T	His445Asp	2
		1355T-C	Leu452Pro	1
		2283G-C	Glu761Asp	1
		508G-T	Val170Phe	1
EMB (11)	embB	916A-G	Met306Val	8
		1216G-A	Gly406Ser	1
		1061A-C	Asp354Ala	1
		1217G-A	Gly406Asp	1
PZA (31)	pncA	281T-G	Phe94Cys	2
		406G-A	Asp136Asn	2
		40T-G	Cys14Gly	1
		304G-C	Ala102Pro	1
		415G-A	Val139Met	1
		226A-C	Thr76Pro	1
		35A-C	Asp12Ala	1
		290G-A	Gly97Asp	1
		22G-A	Asp8Asn	1
		416T-C	Val139Ala	1
		422A-C	Gln141Pro	1
		340A-C	Thr114Pro	1
		464T-G	Val155Gly	1
		490T-C	Ser164Pro	1
		204G-C	Trp68Cys	1
		other	11A-G	2
			390_391dupGG	3
			301dupG	1
			185_186insA	1
			36_37insGCCAGCGAGCCACCCTCGCAGAAGTCG	1
STR (68)	rpsL	128A-G	Lys43Arg	43
		263A-G	Lys88Arg	5
	gid	206G-A	Gly69Asp	1
		517G > T	Gly173*	1
FQs (3)	gyrA	281A-C	Asp94Ala	2
FQs (3)		269C-T	Ala90Val	1
Abbreviation: INH, isoniazid; RIF, rifampicin; EMB, Ethambutol; PZA, pyrazinamide; STR, streptomycin; FQs: fluoroquinolones.

Table 3

Mutations associated with drug-resistance genes in MTB isolates with phenotypic susceptibility to any drug in Finland between 2014 and 2021
Isolates with phenotypic susceptibility to (N)	Gene	Mutation	Amino acid substitution	Frequency
INH (17)	katG	329C-T	Ala110Val	1
EMB (90)	embB	1489C-A	Gln497Lys	1
STR (33)	gid	66C-A	Tyr22*	1
STR (33)	gid	517G-T	Gly173*	1
FQs (98)	gyrA	281A-G	Asp94Gly	1
FQs (98)	gyrA	281A-C	Asp94Ala	1
Abbreviation: INH, isoniazid; EMB, Ethambutol; STR, streptomycin; FQs: fluoroquinolones

Lineage and sub-lineage of 101 MTB isolates

The 101 drug-resistant MTB isolates were classified into four lineages, including lineage 1/Indo-Oceanic, lineage 2/East-Asian, lineage 3/East-African-Indian, and lineage 4/Euro-American (Fig. 1). Across all included drug-resistant MTB isolates, lineage 4 (36/101, 35.6%) and lineage 2 (35/101, 34.6%) were the most prevalent lineages, followed by lineage 1 (16/101, 15.8%) and lineage 3 (14/101, 13.9%). The four lineages were further divided into 21 sub-lineages, which consisted primarily of sub-lineage 2.2.1 (32/101, 31.7%), sub-lineage 3 (13/101, 12.9%), and sub-lineage 4.1.2.1 (12/101, 11.9%). The lineage diversity of drug-resistant MTB isolates was significantly different by regions of birth. Drug-resistant MTB isolates from Finnish-born cases were classified into four lineages, with a dominant sub-lineage 4.1.2.1/Haarlem. Isolates from Asia-born cases were mainly distributed in sub-lineage 1.2.1/EAI2, while most isolates from eastern-Europe-born and sub-Saharan-Africa-born cases were sub-lineage 2.2.1/Beijing and sub-lineage 3/Delhi-CSA, respectively (Table 4).

Table 4

Genetic lineage and sub-lineage of 101 phenotypically drug-resistant MTB isolates in Finland between 2014 and 2021 by region of birth of case
Genetic lineage and sub-lineage ^a	Isolates of Finnish-born cases (N = 29)	Foreign region of birth				Total (N = 101)
Genetic lineage and sub-lineage ^a	Isolates of Finnish-born cases (N = 29)	Asia (N = 24)	Eastern Europe (N = 20)	sub-Saharan Africa (N = 25)	Other / foreign-unknown (N = 3)	Total (N = 101)
Lineage
1	3 (10.3)	10 (41.7)	0	3 (12.0)	0	16 (15.8)
2	7 (24.1)	7 (29.2)	17 (85.0)	3 (12.0)	1 (33.3)	35 (34.7)
3	1 (3.4)	1 (3.4)	0	11 (44.0)	1 (33.3)	14 (13.9)
4	18 (62.1))	6 (20.7)	3 (15.0)	8 (32.0)	1 (33.3)	36 (35.6)
Sub-lineage
1.1.1.1	0	2 (6.9)	0	0	0	2 (2.0)
1.1.2	1 (3.4)	1 (3.4)	0	1 (4.0)	0	3 (3.0)
1.2.1	2 (6.8)	6 (20.7)	0	0	0	8 (7.9)
1.2.2	0	1 (3.4)	0	2 (8.0)	0	3 (3.0)
2.2.1	7 (24.1)	4 (13.8)	17 (85.0)	3 (12.0)	1 (33.3)	32 (31.9)
2.2.1.1	0	3 (10.3)	0	0	0	3 (3.0)
3	0	1 (3.4)	0	11 (44.0)	1 (33.3)	13 (12.8)
3.1.1	1 (3.4)	0	0	0	0	1 (0.9)
4	1 (3.4)	0	0	0	0	1 (0.9)
4.1.1.3	0	0	0	1 (4.0)	0	1 (0.9)
4.1.2.1	9 (31.0)	1 (3.4)	1 (5.0)	0	1 (33.3)	12 (11.9)
4.2.1	3 (10.3)	0	0	0	0	3 (3.0)
4.2.2	0	1 (3.4)	0	3 (12.0)	0	4 (4.0)
4.3.3	0	0	2 (10.0)	2 (8.0)	0	4 (4.0)
4.4	0	2 (6.9)	0	0	0	2 (2.0)
4.5	0	2 (6.9)	0	0	0	2 (2.0)
4.6	0	0	0	1 (4.0)	0	1 (0.9)
4.6.2	0	0	0	1 (4.0)	0	1 (0.9)
4.7	1 (3.4)	0	0	0	0	1 (0.9)
4.8	3 (10.3)	0	0	0	0	3 (3.0)
4.9	1 (3.4)	0	0	0	0	1 (0.9)
^a Genetic lineage and sub-lineage were determined by fast-lineage-caller (version 0.3.1). All values are n (%) unless otherwise indicated.

Transmission clusters

Of 101 drug-resistant MTB isolates, 16 (15.8%) were classified into seven potential transmission clusters, in which the difference between any two isolates was ≤ 12 SNPs. The number of drug-resistant isolates in every potential transmission cluster ranged from two to three. Of potential transmission clusters, four clusters contained isolates from Finland- and foreign-born cases, and one cluster contained isolates from cases born in Asia and sub-Saharan Africa. We also identified one potential transmission cluster that contained only isolates from Finnish-born cases notified between 2020 and 2021 and one that contained only isolates from Eritrea-born cases notified in 2017. Most of the MTB isolates classified into potential transmission clusters were resistant to STR or INH. Among the seven potential transmission clusters, only one cluster was classified as a highly related cluster, in which the difference between the two isolates was ≤ 5 SNPs. The highly related cluster included one pair of MDR MTB isolates (Fig. 2).

This molecular epidemiology study revealed that the drug-resistance profile of MTB isolates was significantly different between Finnish- and foreign-born cases between 2014 and 2021 in Finland. We identified prevalent mutations associated with first- and second-line drug resistance, which provides insights into the treatment of drug-resistant TB. During the study period, the genomic diversity and dominant sub-lineages of drug-resistant MTB isolates were significantly different according to the person’s region of birth. Additionally, this study revealed transmission of drug-resistant MTB isolates was not common between Finnish- and foreign-born persons, suggesting an effective TB management and control, as well as limited close contacts.

Compared to Finnish-born persons with drug-resistant isolates, this study found a significantly higher proportion of MDR-TB in foreign-born persons with drug-resistant isolates (34.7% vs 20.7%), suggesting a potential risk for the native population. This finding is consistent with a previous nationwide study, in which the proportion of MDR-TB in foreign-born cases was significantly higher than that in Finnish-born cases in Finland during 2014-2017¹¹. Similar findings have also been reported in other low-incidence countries with increasing immigrant populations, such as Germany, and UK^9,16. We also identified the most prevalent mutations encoding resistance to INH (katG Ser315Thr), RIF (rpoB Ser450Leu), STR (rpsL Lys43Arg), EMB (embB Met306Val), and FQs (gyrA Asp94Ala). Although we found discrepancies between phenotypic and genotypic resistance results on INH, EMB, STR and FQs, the frequencies were low. Our investigation further emphasises that detection of mutations associated with drug resistance can assist clinicians in diagnosis of DR-TB. Our observation heightened the importance of TB screening for immigrants from high-incidence countries, which has been implemented systematically in some low-incidence settings. Furthermore, systematic follow-up procedures for immigrants from high-incidence countries should be implemented to manage DR-TB.

The 101 drug-resistant MTB isolates included in this study were from four different previously described lineages, namely lineage 1, lineage 2, lineage 3, and lineage 4. Lineage 2 and lineage 4 were prevalent in drug-resistant MTB isolates in Finland. Similar to our results, a WGS-based surveillance on MTB covering 25 European union (EU)/European Economic Area countries reported that the most represented lineages were lineage 4 and lineage 2 among RR or MDR MTB¹⁷. Lineage 4 is most likely of European origin, while lineage 2 originated from eastern Asia¹⁸. The prevalence of lineage 2 in Finland indicated that lineage structure may be driven by human immigration. Moreover, lineage 2 exhibited a higher proportion of MDR-TB and transmissibility than other lineages, suggesting a higher risk to local TB control¹⁹. Additionally, we found differences in the genomic diversity and dominant sub-lineages of drug-resistant MTB between Finnish-born cases and foreign-born cases. Drug-resistant MTB isolates from cases born in Finland, Asia, or sub-Saharan Africa were more genetically diverse (spanning lineage 1 to lineage 4), while those from cases born in eastern Europe consisted only of lineage 2 and lineage 4. Furthermore, lineage 4 was genetically more diverse in cases from Finland, Asia, or sub-Saharan Africa. Combined with our cluster analysis that only four native cases were clustered with foreign-born cases, the transmission of drug-resistant MTB isolates between native cases and cases from Asia or sub-Saharan Africa was not common. Thus, the genetic diversity of lineage 4 in cases born in Asia or sub-Saharan Africa indicated the globally widespread distribution of lineage 4, which was most likely caused by human migration. On the other hand, the low genetic diversity of drug-resistant MTB in cases born in eastern Europe was likely due to limited spread between cases born in eastern Europe and cases from other regions of birth. Another noteworthy finding was that the dominant sub-lineage of drug-resistant MTB from cases born in eastern Europe was sub-lineage 2.2.1 (Beijing) in Finland. This finding is in line with some previous studies conducted in eastern Europe, in which over half of MDR/XDR MTB isolates were classified into lineage 2.2.1^20,21.

Cluster analysis identified seven potential transmission clusters containing a total of 16 drug-resistant MTB isolates (6 from Finnish-born cases and 10 from foreign-born cases) at a threshold of 12 SNPs difference, most of which were formed by lineage 4. The clustering rate (15.8%) of drug-resistant MTB isolates in this low-incidence setting was lower than that (48.5%) in Thailand, which is a high- incidence setting²². Previous studies reported a high clustering rate of MDR isolates in the Czech Republic (29.23%), Spain (48.4%), and EU countries (51.6%)^17,23,24. The lower clustering rate of drug-resistant MTB in Finland may be due to smaller immigrant populations from high-incidence countries in Finland than that in other EU countries. In contrast to our finding that the potential transmission clusters of drug-resistant MTB were mainly composed of lineage 4 (7/16, 43.8%) in Finland, a previous study revealed that lineage 2 exhibited a higher clustering rate than lineage 4 (31.8% vs 25%) in Sweden²⁵. This inconsistency is likely due to differences in immigration structures between the two countries. Although we identified seven potential transmission clusters, only four Finnish-born cases were clustered with foreign-born cases, suggesting a limited transmission of drug-resistant MTB between Finnish- and foreign-born cases. Furthermore, one of the potential transmission clusters was genetically highly related (≤ 5 SNPs difference), which contained one Finnish-born case and one case born in eastern Europe. The two isolates with a below 5 SNPs difference were sub-lineage 2.2.1 and MDR. Consistent with our finding, a study using MIRU-VNTR typing found that MTB transmission between the foreign and Finnish-born population was uncommon between 2014 and 2017¹¹. Compared to this study, previous molecular epidemiology studies found more transmission events of MTB between native populations and immigrants in Switzerland, Germany, Sweden, UK and France^8,9,16,25,26. The reasons may be partly attributed to the lack of systematic follow-up procedures for foreign-born cases in these low-incidence settings. Our findings showed that WGS analysis may provide further insights into transmission dynamics of drug-resistant MTB in low-incidence settings with a high proportion of immigrants, especially when social contact tracing is difficult to perform²⁷. Interpretation of small SNP differences needs to be careful if no epidemiological links are identified, since infections may be distantly acquired and other factors, such as clinical disease manifestation, affect the assessment of transmission²⁸.

Our study has some limitations. First, this study lacked traditional epidemiological contact data and immigration routes, which are resource-consuming to acquire and were not the focus here. We investigated the molecular epidemiological characteristics and seeked potential transmission events of drug-resistant MTB between Finnish- and foreign-born cases based on WGS data. Another limitation of our study is the definition of transmission clusters. Many previous studies used different thresholds of SNPs difference (5–12 SNPs) to determine a genetic cluster. Our WGS data spanned from 2014 to 2021. To provide as much insight as possible into transmission dynamics of drug-resistant MTB, we used a threshold of 12 SNPs to determine a potential related cluster, which means a potential transmission event. In addition, we also used a threshold of 5 SNPs to determine a highly related cluster, which means a recent transmission.

In conclusion, the genetic diversity of drug-resistant MTB isolates observed in Finland is more complex than expected. The genetic diversity of drug-resistant MTB isolates varied significantly according to region of birth of cases, suggesting most cases were infected with drug-resistant MTB in the region of birth. Surveillance of drug-resistant MTB isolates from foreign-born cases is necessary for optimal local TB management. Although transmission of drug-resistant MTB isolates between Finnish- and foreign-born cases was less widespread, the risk of drug-resistant MTB for native population is not negligible. Understanding the transmission dynamics and geographical dispersion of genetic lineages of drug-resistant MTB isolates is of great importance to evaluate and optimise drug-resistant TB control measures in low-incidence countries with immigration from high-incidence countries.

Ethics

The data used for the study was collected as part of the infectious disease surveillance and control, defined by national legislation and ethical approval was therefore not necessary.

Funding

Authors would like to thank the funding by Helsinki University Hospital Funds (projects TYH2018322, TYH2018322 and M1023TK001), and the Jane and Aatos Erkko Foundation.

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Competing interests: The authors have no conflicts of interests to disclose.

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Transmission of drug-resistant Mycobacterium tuberculosis isolates between Finnish- and foreign-born patients, 2014-2021: a molecular epidemiological study

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