Pyrazinamide Resistance and pncA Mutation Profiles in Multidrug Resistant Mycobacterium Tuberculosis

Figures & data

Table 1 Demographic Characteristics of Patients and Genotyping of Clinical Strains Resistant and Susceptible to PZA

	PZA Susceptibility Test		p value
	Resistant	Sensitive
Means of age	42.36	42.71	0.927^c
Gender^a			0.335
Male	12	38
Female	10	16
Treatment history^b			0.758
New cases	5	14
Previously treated	18	42
Genotyping			1.000
Beijing family	22	57
Non-Beijing	1	2

Notes: ^aSix patients without gender documentation. ^bThree patients without treatment history documentation. ^cCalculated by t-test.

Table 2 Characteristics of pncA Mutation, PZA Susceptibility and Genotyping in pncA Mutated MDR Strains

Mutation Site (Nucleotide Position)	Nucleotide Change	Amino Acid Position	Amino Acid Replacement	Activity of PZAse	No. of Strains	Pyrazinamide Susceptibility^a	Beijing/non-Beijing Genotype	Documented in WHO Guide
3	G→A	1	Met→Ile	20.5%	1	Susceptible	Beijing	No
20	T→G	7	Val→Gly	9.79%	1	Resistant^b	Beijing	Yes, moderate
35	A→C	12	Asp→Ala	0.0%	2	One resistant^b, one susceptible	Beijing	Yes
35	A→G	12	Asp→Gly	0.0%	1	Susceptible	Beijing	Yes, minimal
40	T→C	14	Cys→Arg	14.6%	1	Resistant^b	Beijing	Yes
56	T→C	19	Leu→Pro	8.0%	1	Susceptible	Beijing	Yes
72–506	435bp fragment deletion	24–169	Frameshift	0.0%	1	Susceptible	Beijing	No
79–92	del CTGGCCC GCGCCAT	27–31	Frameshift	F	1	Resistant^b	Beijing	No
80	T→G	27	Leu→Arg	0.0%	1	Susceptible	Beijing	No
83	C→A	28	Ala→Asp	21.0%	1	Susceptible	Beijing	No
104–105	TG→CC	35	Leu→Pro	29.0%	1	Resistant	Beijing	No
146	A→C	49	Asp→Ala	0.0%	1	Resistant^b	Beijing	Yes
161–162	CG→GC	54	Pro→Arg	6.1%	1	Resistant^b	Beijing	No
170	A→G	57	His→Arg	0.0%	1	Resistant	Beijing	Yes
171	C→A	57	His→Gln	0.0%	1	Resistant	Non-Beijing	Yes
172	T→G	58	Phe→Val	0.0%	1	Susceptible	Beijing	No
195	C→T	65	Synonymous	90.5%	1	Resistant	Beijing	No
203	G→A	68	Trp→Stop	35.2%	1	Susceptible	Beijing	No
203	G→C	68	Trp→Ser	25.3%	1	Susceptible	Beijing	No
203	Ins GT	68	Frameshift	2.0%	1	Susceptible	Beijing	No
212	A→G	71	His→Arg	0.0%	1	Susceptible	Beijing	Yes, minimal
226	A→C	76	Thr→Pro	32.5%	2	One resistant, one susceptible	Beijing	Yes
227	C→T	76	Thr→Ile	0.0%	1	Susceptible	Beijing	No
246	del T	82	Frameshift	0.0%	1	Susceptible	Beijing	No
269	T→C	90	Ile→Thr	17.1%	1	Susceptible	Beijing	No
305	C→T	102	Ala→Val	20.0%	1	Susceptible	Beijing	No
308	A→G	103	Tyr→Cys	12.2%	1	Resistant^b	Beijing	No
347	T→G	116	Leu→Arg	0.0%	1	Susceptible	Beijing	No
359	T→G	120	Leu→Arg	0.0%	1	Resistant^b	Beijing	No
398	T→C	133	Ile→Thr	9.8%	1	Resistant	Beijing	Yes, minimal
404	C→A	135	Thr→Asn	12.9%	1	Susceptible	Beijing	No
416	T→C	139	Val→Ala	0.0%	1	Susceptible	Beijing	No
418	Ins TG	140	Frameshift	0.0%	4	Resistant^c	Beijing	No
463	G→A	155	Val→Met	7.0%	1	Susceptible	Beijing	No
464	T→C	155	Val→Ala	21.8%	1	Susceptible	Beijing	No
467	T→A	156	Leu→Gln	19.9%	1	Susceptible	Beijing	No
478	A→C	160	Thr→Pro	0.0%	1	Resistant^b	Beijing	No
Total					42

Notes: ^aResults of pyrazinamide susceptibility testing based on the critical concentration 100 mg/L. ^bResistant to 300mg/L PZA. ^cAll four strains resistant to 100 mg/L PZA and two of them resistant to 300 mg/L PZA. Confidence grading of pncA mutations in WHO GUIDE were classified as high, moderate and minimal.⁵⁰

Abbreviation: F, fail to express the protein.

Table 3 Concordance of pncA Mutation Detection and PZA Susceptibility by MGIT 960

PZA Suscptibility		pncA Genotype		Total Strains	P value	Sensitivity (%)	Specificity (%)
		Mutant	Wildtype
100mg/L					0.001	82.6	61.0
Resistant		19	4	23
Susceptible		23	36	59
300mg/L					0.009	84.6	55.1
Resistant		11	2	13
Susceptible		31	38	69

Figure 1 Pyrazinamidase (PZAse) activity of strains. Red: susceptible to pyrazinamide (PZA) at 100 mg/L; Blue: resistant to PZA at 100 mg/L; Green: resistant to PZA at 300 mg/L. *Mutation type with different phenotype.

World Health Organization. The use of next-generation sequencing technologies for the detection of mutations associated with drug resistance in Mycobacterium tuberculosis complex: technical guide. World Health Organization; 2018.

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