微生物学报

Test number	Level	NH₄⁺-N removal efficiency/%
1	25	7.0	15	180	71.55
2	25	7.0	20	160	78.62
3	25	6.0	15	160	71.41
4	25	7.0	15	140	70.13
5	25	8.0	15	160	79.59
6	25	7.0	10	160	69.85
7	30	6.0	20	160	81.44
8	30	6.0	15	180	80.86
9	30	7.0	20	180	89.17
10	30	6.0	15	140	79.59
11	30	6.0	10	160	76.55
12	30	8.0	15	180	82.81
13	30	7.0	10	140	79.54
14	30	7.0	15	160	96.87
15	30	7.0	15	160	99.97
16	30	7.0	15	160	94.03
17	30	7.0	15	160	92.88
18	30	7.0	15	160	99.01
19	30	8.0	10	160	84.42
20	30	7.0	20	140	78.43
21	30	8.0	15	140	80.94
22	30	7.0	10	180	80.52
23	30	8.0	20	160	83.71
24	35	7.0	15	180	82.72
25	35	6.0	15	160	80.00
26	35	7.0	15	140	86.61
27	35	8.0	15	160	85.20
28	35	7.0	20	160	80.43
29	35	7.0	10	160	83.94

Test number	Level	NH₄⁺-N removal efficiency/%
1	25	7.0	15	180	71.55
2	25	7.0	20	160	78.62
3	25	6.0	15	160	71.41
4	25	7.0	15	140	70.13
5	25	8.0	15	160	79.59
6	25	7.0	10	160	69.85
7	30	6.0	20	160	81.44
8	30	6.0	15	180	80.86
9	30	7.0	20	180	89.17
10	30	6.0	15	140	79.59
11	30	6.0	10	160	76.55
12	30	8.0	15	180	82.81
13	30	7.0	10	140	79.54
14	30	7.0	15	160	96.87
15	30	7.0	15	160	99.97
16	30	7.0	15	160	94.03
17	30	7.0	15	160	92.88
18	30	7.0	15	160	99.01
19	30	8.0	10	160	84.42
20	30	7.0	20	140	78.43
21	30	8.0	15	140	80.94
22	30	7.0	10	180	80.52
23	30	8.0	20	160	83.71
24	35	7.0	15	180	82.72
25	35	6.0	15	160	80.00
26	35	7.0	15	140	86.61
27	35	8.0	15	160	85.20
28	35	7.0	20	160	80.43
29	35	7.0	10	160	83.94

Source	Sum of squares	df	Mean square	F-value	P-value	Significant
Model	1 664.76	14	118.91	15.51	<0.000 1	**
A-temperature	277.99	1	277.99	36.25	<0.000 1	**
B-pH	63.88	1	63.88	8.33	0.012 0	*
C-C/N	23.98	1	23.98	3.13	0.098 7	-
D-shaking speed	11.06	1	11.06	1.44	0.249 7	-
AB	2.23	1	2.23	0.29	0.598 2	-
AC	37.69	1	37.69	4.92	0.043 7	*
AD	7.03	1	7.03	0.92	0.354 5	-
BC	7.81	1	7.81	1.02	0.329 9	-
BD	0.53	1	0.53	0.07	0.796 4	-
CD	23.83	1	23.83	3.11	0.099 8	-
A²	736.20	1	736.20	96.01	<0.000 1	**
B²	360.13	1	360.13	46.97	<0.000 1	**
C²	348.39	1	348.39	45.43	<0.000 1	**
D²	405.52	1	405.52	52.89	<0.000 1	**
Residual	107.35	14	7.67	-	-	-
Lack of fit	69.68	10	6.97	0.74	0.682 6	-
Pure error	37.67	4	9.42	-	-	-
Cor total	1 772.11	28	-	-	-	-

Source	Sum of squares	df	Mean square	F-value	P-value	Significant
Model	1 664.76	14	118.91	15.51	<0.000 1	**
A-temperature	277.99	1	277.99	36.25	<0.000 1	**
B-pH	63.88	1	63.88	8.33	0.012 0	*
C-C/N	23.98	1	23.98	3.13	0.098 7	-
D-shaking speed	11.06	1	11.06	1.44	0.249 7	-
AB	2.23	1	2.23	0.29	0.598 2	-
AC	37.69	1	37.69	4.92	0.043 7	*
AD	7.03	1	7.03	0.92	0.354 5	-
BC	7.81	1	7.81	1.02	0.329 9	-
BD	0.53	1	0.53	0.07	0.796 4	-
CD	23.83	1	23.83	3.11	0.099 8	-
A²	736.20	1	736.20	96.01	<0.000 1	**
B²	360.13	1	360.13	46.97	<0.000 1	**
C²	348.39	1	348.39	45.43	<0.000 1	**
D²	405.52	1	405.52	52.89	<0.000 1	**
Residual	107.35	14	7.67	-	-	-
Lack of fit	69.68	10	6.97	0.74	0.682 6	-
Pure error	37.67	4	9.42	-	-	-
Cor total	1 772.11	28	-	-	-	-

Nitrogen source	Initial nitrogen concentration/(mg/L)	c(TN)/(mg/L)	Cell-N	Gaseous-N (N₂+N₂O)
NH₄⁺-N	134.07±0.27	10.77±0.17	0.72±0.36	116.87±4.85	64.09±0.21	41.89±1.71	35.84	41.97	35.91
NO₃^--N	2.98±1.08	98.35±3.17	0.22±0.04	108.21±6.45	81.18±0.42	48.52±0.39	44.84	22.62	20.90
NO₂^--N	11.57±2.24	0.00±0.00	91.81±6.28	115.77±0.27	83.45±0.32	47.84±1.73	41.32	21.98	18.99

Nitrogen source	Initial nitrogen concentration/(mg/L)	c(TN)/(mg/L)	Cell-N	Gaseous-N (N₂+N₂O)
NH₄⁺-N	134.07±0.27	10.77±0.17	0.72±0.36	116.87±4.85	64.09±0.21	41.89±1.71	35.84	41.97	35.91
NO₃^--N	2.98±1.08	98.35±3.17	0.22±0.04	108.21±6.45	81.18±0.42	48.52±0.39	44.84	22.62	20.90
NO₂^--N	11.57±2.24	0.00±0.00	91.81±6.28	115.77±0.27	83.45±0.32	47.84±1.73	41.32	21.98	18.99

Bacterial genus name	NH₄⁺-N removal efficiency/%	NO₃^--N removal efficiency/%	NO₂^--N removal efficiency/%	N₂ production	Genes related to nitrogen metabolism	References
Acinetobacteroleivorans AHP123	97.93	-	-	Heterotrophic nitrification: 12.4 mg/L Aerobic denitrification: 18.33 mg/L	nar, amt, glnK, gltB, gdhA, gam, glnA, ncd2, ncd1, nirB, nasA	[7]
Klebsiella sp. TSH15	92.55	91.43	-	-	amt, glnA, gdhA, nirB, narG, napA, nasA, nirK/S, norB, nosZ	[11]
Candida boidinii L21	96.00	98.00	-	-	-	[12]
Pseudomonas aeruginosa WS-03	95.81	94.00	100.00	-	narB, glnA, gdhA, nirB, nirA, napA, gltB	[19]
Acinetobacter ZQ‑A1	84.84	87.13	92.63	-	-	[35]
Pseudomonas sp. Y-5	97.26	99.34	95.81	-	-	[36]
Pseudomonas mendocina TJPU04	98.00	93.00	100.00	-	-	[37]
Klebsiella sp. WH-E	99.80	81.54	80.00	With NH₄⁺-N as the sole nitrogen source: 35.91% With NO₂^--N as the sole nitrogen source: 20.90% With NO₃^--N as the sole nitrogen source: 18.99%	glnB, glnL, glnG, gdhA, norVWR, narGHI, nasBC, nirBD

Bacterial genus name	NH₄⁺-N removal efficiency/%	NO₃^--N removal efficiency/%	NO₂^--N removal efficiency/%	N₂ production	Genes related to nitrogen metabolism	References
Acinetobacteroleivorans AHP123	97.93	-	-	Heterotrophic nitrification: 12.4 mg/L Aerobic denitrification: 18.33 mg/L	nar, amt, glnK, gltB, gdhA, gam, glnA, ncd2, ncd1, nirB, nasA	[7]
Klebsiella sp. TSH15	92.55	91.43	-	-	amt, glnA, gdhA, nirB, narG, napA, nasA, nirK/S, norB, nosZ	[11]
Candida boidinii L21	96.00	98.00	-	-	-	[12]
Pseudomonas aeruginosa WS-03	95.81	94.00	100.00	-	narB, glnA, gdhA, nirB, nirA, napA, gltB	[19]
Acinetobacter ZQ‑A1	84.84	87.13	92.63	-	-	[35]
Pseudomonas sp. Y-5	97.26	99.34	95.81	-	-	[36]
Pseudomonas mendocina TJPU04	98.00	93.00	100.00	-	-	[37]
Klebsiella sp. WH-E	99.80	81.54	80.00	With NH₄⁺-N as the sole nitrogen source: 35.91% With NO₂^--N as the sole nitrogen source: 20.90% With NO₃^--N as the sole nitrogen source: 18.99%	glnB, glnL, glnG, gdhA, norVWR, narGHI, nasBC, nirBD

克雷伯氏菌属(Klebsiella sp.) WH-E的筛选及脱氮机制

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巩璐琳 ¹ , 马咏琪 ¹ , 张爱文 ² , 张艳丽 ³ , 孙旭春 ⁴ , 张引弟 ¹ , 漆文瑞 ¹ , 薛媛 ¹ , 孙丽坤 ¹

微生物学报 | 研究报告 2026,66(5): 2174-2190

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微生物学报 | 研究报告 2026, 66(5): 2174-2190

克雷伯氏菌属(Klebsiella sp.) WH-E的筛选及脱氮机制

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巩璐琳¹, 马咏琪¹, 张爱文², 张艳丽³, 孙旭春⁴, 张引弟¹, 漆文瑞¹, 薛媛¹, 孙丽坤¹

作者信息

^1.甘肃农业大学动物科学技术学院，甘肃兰州

^2.甘肃省畜牧技术推广总站，甘肃兰州

^3.平凉市农产品质量安全与检验检测中心，甘肃平凉

^4.临夏回族自治州畜牧技术推广站，甘肃临夏

Screening and denitrification mechanisms of Klebsiella sp. WH-E

Lulin GONG¹, Yongqi MA¹, Aiwen ZHANG², Yanli ZHANG³, Xuchun SUN⁴, Yindi ZHANG¹, Wenrui QI¹, Yuan XUE¹, Likun SUN¹

Affiliations

^1.College of Animal Science and Technology, Gansu Agricultural University, Lanzhou, Gansu, China

^2.Gansu Provincial Animal Husbandry Technology Promotion Station, Lanzhou, Gansu, China

^3.Pingliang Inspection and Testing Center of Agricultural Product Quality and Safety, Pingliang, Gansu, China

^4.Animal Husbandry Technology Extension Station of Linxia Hui Autonomous Prefecture, Linxia, Gansu, China

出版时间: 2026-05-04 doi: 10.13343/j.cnki.wsxb.20250722

文章导航

摘要

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目的畜禽养殖污水排放量激增，环境污染问题日益凸显。筛选高效异养硝化好氧反硝化菌株并探究其脱氮机制，对解决养殖污水氮素污染问题具有重要的理论与实践意义。方法从猪场活性污泥中筛选一株异养硝化-好氧反硝化(heterotrophic nitrification-aerobic denitrification, HN-AD)能力优异的菌株，采用响应面法优化其培养条件；通过测定菌株对单一和混合氮源的利用效率，借助氮平衡分析方法，评估其对无机氮的转化能力；结合气相色谱法测定N₂O、N₂以验证反硝化过程的完整性；结合全基因组分析解析菌株WH-E的脱氮途径及机制。结果成功分离出一株克雷伯氏菌属(Klebsiella sp.) WH-E，该菌株具有优异的异养硝化好氧反硝化性能。经优化试验确定其最适生长参数为：碳源为柠檬酸钠，温度为34.18 ℃，初始pH为7.1，C/N为14.53，摇床转速为159.59 r/min。菌株以氨氮、硝氮、亚硝氮为单一氮源培养时氮素去除率分别为99.80%、81.54%、80.00%；以氨氮为唯一氮源时，35.84%和35.91%的氮分别被转化为细胞氮和气态氮。以氨氮和硝氮为混合氮源培养时氮素去除率均为100.00%；以氨氮和亚硝氮为混合氮源培养时，氨氮去除率为100.00%，亚硝氮去除率为91.97%。全基因组测序分析发现glnB、norVWR、narGHI、nasBC、nirBD等多个与氮代谢相关的功能基因。结论 Klebsiella sp. WH-E有3种氮代谢途径，包括氨同化途径、硝化反硝化途径和硝酸盐同化与异化途径。本研究证实了HN-AD菌株在实际养殖废水脱氮处理中的应用潜力，并为其工程化实施提供了理论支撑。

关键词

异养硝化好氧反硝化菌 / 克雷伯氏菌属(Klebsiella sp.) WH-E / 脱氮性能 / 氮代谢 / 全基因组

Abstract

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Objective The rapid increase in wastewater discharge from animal husbandry has caused severe environmental pollution. Identifying efficient heterotrophic nitrifying-aerobic denitrifying bacteria and investigating their denitrification mechanisms are of great theoretical and practical importance for mitigating nitrogen pollution in the wastewater. Methods A strain exceling in heterotrophic nitrification-aerobic denitrification (HN-AD) was isolated and from activated sludge in pig farms. Culture conditions were optimized by response surface methodology. We evaluated the inorganic nitrogen-transforming capacity of the strain by assessing its utilization efficiency of single and mixed nitrogen sources and through nitrogen balance analysis. The completeness of the denitrification process was confirmed via gas chromatographic measurements of N₂ and N₂O. Finally, the nitrogen removal pathways and underlying mechanisms were elucidated through whole-genome analysis. Results The successfully isolated strain Klebsiella sp. WH-E exhibited excellent HN-AD capabilities. The growth conditions of the strain were optimized as follows: sodium citrate as the carbon source, 34.18 °C, initial pH 7.1, a C/N ratio of 14.53, and a shaking speed of 159.59 r/min. When the strain was cultured with ammonium, nitrate, or nitrite as the sole nitrogen source, the nitrogen removal rates were 99.80%, 81.54%, and 80.00%, respectively. Furthermore, when ammonium was the sole nitrogen source, 35.84% and 35.91% of nitrogen were converted into cellular nitrogen and gaseous nitrogen, respectively. When ammonia nitrogen was combined with nitrate nitrogen as mixed nitrogen sources, the nitrogen removal rate was 100.00%; When ammonia nitrogen was combined with nitrite nitrogen as mixed nitrogen sources, the ammonia nitrogen removal rate was 100.00%, and the nitrite nitrogen removal rate was 91.97%, respectively. Whole-genome sequencing identified several nitrogen metabolism-related functional genes, including glnB, norVWR, narGHI, nasBC, and nirBD. Conclusion Klebsiella sp. WH-E possesses three nitrogen metabolism pathways: ammonium assimilation, nitrification-denitrification, and nitrate assimilation and dissimilation. This study confirms the applicability of Klebsiella sp. WH-E for nitrogen removal from full-scale piggery wastewater and establishes a solid theoretical foundation for its engineering applications.

Key words

heterotrophic nitrifying-aerobic denitrifying bacteria / Klebsiella sp. WH-E / denitrification performance / nitrogen metabolism / whole genome

引用本文

巩璐琳, 马咏琪, 张爱文, 张艳丽, 孙旭春, 张引弟, 漆文瑞, 薛媛, 孙丽坤. 克雷伯氏菌属(Klebsiella sp.) WH-E的筛选及脱氮机制. 微生物学报, 2026 , 66 (5) : 2174 -2190 . DOI: 10.13343/j.cnki.wsxb.20250722

Lulin GONG, Yongqi MA, Aiwen ZHANG, Yanli ZHANG, Xuchun SUN, Yindi ZHANG, Wenrui QI, Yuan XUE, Likun SUN. Screening and denitrification mechanisms of Klebsiella sp. WH-E[J]. Acta Microbiologica Sinica, 2026 , 66 (5) : 2174 -2190 . DOI: 10.13343/j.cnki.wsxb.20250722

正文

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畜禽的集约化发展导致污水排放量急剧增加，给生态环境造成了巨大压力。《第二次全国污染源普查公报》指出，畜禽养殖废水中氨氮(ammonium nitrogen, NH₄⁺-N)的排放量为7.50×10⁴ t^[1]。高浓度的有机氮不仅会影响水体的自净能力，还会危及人或动物的健康^[2]。因此，去除水中的有机氮、维护水体健康和可持续性，已成为重要的研究方向。

生物脱氮因脱氮效率高、成本低且无二次污染，成为当前养殖废水脱氮的核心技术^[3]，其关键在于利用微生物转化实现氮素去除。异养硝化好氧反硝化(heterotrophic nitrifying-aerobic denitrifying, HN-AD)菌的发现为生物脱氮提供了新思路，此类微生物因其硝化和反硝化在时间和空间上能够同步进行而受到广泛关注，且在高氨氮废水的处理中体现出显著的环境和经济价值^[4]。目前已发现包括克雷伯氏菌属(Klebsiella)^[5]、假单胞菌属(Pseudomonas)^[6]和不动杆菌属(Acinetobacter)^[7]在内的多种菌株具有HN-AD能力。其氮代谢途径主要有2种：(1) 完全硝化反硝化途径[NH₄⁺-N→氢氧化胺(hydroxy lamine, NH₂OH)→硝酸盐氮/亚硝酸盐氮(nitrate nitrogen/nitrite nitrogen, NO₂^--N/NO₃^--N)→氮气(nitrogen gas, N₂)]，如菌株铜绿假单胞菌(Pseudomonas aeruginosa)^[8]；(2) NH₂OH还原脱氮途径[NH₄⁺-N→NH₂OH→一氧化二氮(nitrous oxide, N₂O)/N₂]，如菌株盐单胞菌属(Halomonas sp.) DN3^[9]。近年来，研究发现Klebsiella. sp. TN-10^[10]对硝酸盐和亚硝酸盐的去除效率分别为95.44%和99.87%；Klebsiella sp. TSH15^[11]和Candida boidinii L21^[12]等菌株的全基因组分析也证实其具有氨同化和同化/异化硝酸盐还原代谢途径。然而，针对异养硝化好氧反硝化菌对畜禽养殖废水中不同氮源的高效脱氮菌株的研究，以及对其多重脱氮机制的全面解析仍存在不足，特别是对脱氮终产物气态氮的定量验证研究较少。

与Klebsiella sp. TN-10^[10]和Klebsiella sp. TSH15^[11]等仅开展单因素试验或初步基因组分析的菌株研究相比，本研究通过筛选脱氮性能优异的HN-AD菌并采用响应面法优化其培养条件，定量分析各类无机氮的浓度变化及N₂的生成量来评估脱氮能力，同时结合全基因组数据分析其脱氮途径，明确并定量解析了菌株在多重氮源条件下的脱氮性能与气态氮转化途径。本研究的创新点在于结合气态氮定量和全基因组注释阐明多条氮代谢机制，为实际养殖废水脱氮提供理论依据和菌种资源。

1 材料与方法

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1.1 样品来源

污水样本采自甘肃省三利众合农业开发有限公司的猪场。

1.2 培养基

富集(LB)培养基，参考马咏琪等^[13]的研究。

硝化培养基(basal medium, BM)、反硝化培养基(denitrification medium, DM)、同步异养硝化好氧反硝化培养基(heterotrophic nitrification and aerobic denitrification medium, HM)，均参考马咏琪^[14]的研究。

后续所有试验均设置3个重复，并以不接种菌液的培养基作为对照。

1.3 菌株的筛选与鉴定

参考马咏琪^[14]的研究，筛选并鉴定具备高效脱氮能力的菌株，成功获得一株性能优异的菌株，命名为WH-E，用于后续深入探究。

1.4 菌株的脱氮性能测定

将菌液按1%的体积分数分别接入BM与DM培养基后，在碳源为柠檬酸钠、pH为7.0、C/N为15、温度为30 °C、转速为140 r/min的培养条件下培养48 h。每隔12 h取样，测定OD₆₀₀、NO₃^--N、NO₂^--N、NH₄⁺-N浓度，并计算氮素去除率。测定方法和计算公式参考马咏琪^[14]和Zhang等^[15]的研究。

1.5 菌株培养条件优化

1.5.1 单因素试验

涉及5种常见环境因子，包括碳源(葡萄糖、柠檬酸钠、丁二酸钠、蔗糖、乙酸钠)、C/N (5-25)、pH (5.0-10.0)、温度(20-40 ℃)、转速(100-180 r/min)。鉴于畜禽养殖废水中NH₄⁺-N是主要的污染形态，本研究选择以NH₄⁺-N作为目标氮源进行条件优化。将菌悬液接入BM培养基中，每隔12 h取样，测定OD₆₀₀、NH₄⁺-N、NO₂^--N、NO₃^--N浓度，并计算NH₄⁺-N去除率。测定方法与计算公式同1.4节。

1.5.2 响应面优化

采用Box-Behnken构建以柠檬酸钠为碳源的四因素三水平响应面模型，考察因素与范围为C/N比15-25、初始pH 7.0-9.0、温度25-35 °C、转速140-180 r/min。在250 mL锥形瓶中培养48 h，分别于试验起始与结束时取样，检测NH₄⁺-N变化并计算其去除率。

1.6 氮代谢特性分析

将对数生长期的WH-E菌悬液分别接入BM、DM及HM培养基中，在碳源为柠檬酸钠、pH为7.0、C/N为15、温度为30 °C、转速为180 r/min的最佳培养条件下培养48 h。每隔12 h取样，测定OD₆₀₀后4 000 r/min离心10 min，并检测上清中OD₆₀₀、NO₃^--N、NH₄⁺-N、NO₂^--N、羟胺(hydroxy lamine, NH₂OH-N)、总氮(total nitrogen, TN)、细胞氮(cellular nitrogen, Cell-N)、总溶解态氮(total dissolved nitrogen, DTN)浓度，同时计算氮素去除率，计算公式同1.4节。OD₆₀₀、NO₃^--N、NH₄⁺-N、NO₂^--N测定方法同1.4节。NH₂OH-N采用间接分光光度法测定，TN、Cell-N、DTN采用碱性过硫酸钾紫外分光光度法。Cell-N浓度计算公式参考Liu等^[16]，如公式(1)所示。

C=C_TN-C_DTN(1)

式中：C为细胞氮浓度(mg/L)，C_TN为总氮浓度(mg/L)，C_DTN为总溶解态氮浓度(mg/L)。

1.7 氮代谢途径中气体产物检测

分别以NH₄⁺-N、NO₃^--N与NO₂^--N作为唯一氮源，将WH-E菌悬液接入500 mL厌氧血清瓶中进行培养，培养基体积为200 mL。使用纯氧置换法排净瓶内气体并密封，在最佳培养条件下培养48 h。培养结束时借助气密性注射器抽取100 mL顶空气体于铝箔集气袋。气体收集装置如图1所示。最后采用配备电子捕获检测器(electron capture detector, ECD)的气相色谱设备测定N₂O，配备热导检测器(thermal conductivity detector, TCD)的气相色谱设备测定N₂、氧气(oxygen, O₂)，实际进样时使用1 000 μL微量进样器自气体密封袋中取样测定。

1.8 全基因组测试

将菌液接种至BM培养基中培养至对数生长期，于4 °C、1 000 r/min离心10 min收集菌体，经PBS缓冲液漂洗3次后，委托上海美吉生物医药科技有限公司完成全基因组测序工作。采用Illumina第二代测序平台和PacBio第三代测序技术联合策略构建菌株基因组文库，并实施Illumina高通量测序。对获得的序列进行基因注释与预测，基于GO (https://geneontology.org/)、KEGG (https://www.kegg.jp/)、COG (http://www.ncbi.nlm.nih.gov/COG/)、NR (ftp://ftp.ncbi.nih.gov/pub/nrdb/)、Pfam (http://pfam.xfam.org/)、TCDB (http://www.tcdb.org/)和Swiss-Prot (https://www.uniprot.org/)等数据库对编码序列进行基因功能注释。

1.9 数据分析方法

使用Excel 2019与SPSS 27.0分别进行数据整理与统计分析，结果用平均值±标准差表示，P<0.05表示差异显著，所有图表均用Origin 2022软件绘制。

2 结果与分析

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2.1 菌株的分离与鉴定

在平板上，菌株WH-E菌落呈现淡黄色，微隆起，边缘整齐(图2A)。经革兰氏染色鉴定，该菌株属于革兰氏阴性菌。通过扫描电子显微镜观察，细胞形态为棒杆状，大小约为1.0 μm×(1.5-2.0) μm (图2B)。如图2C所示，菌株WH-E应归属于克雷伯氏菌属(Klebsiella sp.)，其16S rRNA基因序列与该菌株(GenBank登录号为OR816101)的相似度达到99%。

2.2 单一氮源时菌株的脱氮性能

如图3所示，在以(NH₄)₂SO₄为唯一氮源时，WH-E在12-36 h进入对数生长期，培养结束时，有90.59%的NH₄⁺-N被去除。以NaNO₃为唯一氮源时，菌株OD₆₀₀在48 h时达到峰值，同时NO₃^--N的去除率为86.65%。在以NO₂^--N为唯一氮源的体系中，菌株在12-24 h进入快速生长期，并于48 h NO₂^--N被利用92.58%。

2.3 Klebsiella sp. WH-E培养条件优化

2.3.1 单因素分析

如图4-6所示，菌株利用柠檬酸钠的效果显著高于其余碳源(P<0.05)，其OD₆₀₀峰值达0.9，且NH₄⁺-N去除效率最高，为80.47%。以柠檬酸钠为碳源时，NO₃^--N和NO₂^--N的积累量分别为7.30 mg/L和1.04 mg/L，均低于其他碳源条件，表明该菌株的最优碳源为柠檬酸钠。当C/N设定为15、20和25时，培养48 h时对应的菌株OD₆₀₀值和NH₄⁺-N去除率均显著高于C/N为5和10 (P<0.05)；NO₃^--N的积累量分别为0.50、0.63、0.65 mg/L，NO₂^--N的积累量分别为1.65、2.90、2.79 mg/L。因此，菌株WH-E的最佳C/N为15。在30 °C条件下菌株生长速度和NH₄⁺-N去除率显著优于其他温度条件(P<0.05)，48 h时OD₆₀₀为1.4，NH₄⁺-N去除率为86.11%，NO₃^--N和NO₂^--N的积累量分别为1.17 mg/L和1.40 mg/L，表明该菌的最适温度为30 °C。转速试验表明，提高转速有利于菌株生长与脱氮，在140、160、180 r/min时，48 h时菌株OD₆₀₀分别为1.4、1.3和1.3，NH₄⁺-N去除率分别为86.12%、86.83%和88.53%，显著优于100 r/min和120 r/min (P<0.05)；NO₃^--N的积累量分别为0.53、0.63、0.72 mg/L，NO₂^--N的积累量分别为1.07、1.57、1.52 mg/L。因此，最佳摇床转速为180 r/min。pH范围为7.0-9.0时OD₆₀₀≈1.0，NH₄⁺-N去除率均维持在较高水平。pH 7.0时NH₄⁺-N去除率达到最高值86.11%。NO₃^--N和NO₂^--N的积累量分别为0.27 mg/L和1.08 mg/L。因此，WH-E的最佳pH为7.0。为便于后续响应面试验有一个能够有效捕捉因素交互作用与潜在非线性效应的优化区间，选取C/N为15-25、温度为25-35 °C、转速为140-180 r/min、pH为7.0-9.0作为考察范围。

2.3.2 响应面分析

依据单因素试验结果，基于Box-Behnken设计对C/N、初始pH、温度、转速4个关键变量进一步优化(表1)。构建以NH₄⁺-N去除率为响应值的二次回归模型，方程如下：

Y(%)=96.55+4.81A+2.31B+1.41C+0.96D-0.75AB-3.07AC-1.33AD-1.40BC+0.36BD+2.44CD-10.65A²-7.45B²-7.33C²-7.91D²，R²为0.939 4。

方差分析结果如表2所示，各因素对菌株脱氮性能的影响程度依次为：温度>pH>C/N>转速。由图7可知，不同环境因子表现出显著的交互效应。模型模拟得出最佳培养参数：在以柠檬酸钠为有机碳源，温度为34.18 °C，初始pH为7.1，C/N为14.53，转速为159.59 r/min的条件下，NH₄⁺-N去除率的理论预测值是93.29%。通过3组平行验证试验，实际去除率为93.30%，与理论值偏差为0.10%，充分证实所构建的模型具备高度的精确性。

2.4 Klebsiellasp. WH-E氮代谢特性

结合菌株对氮平衡分析(表3)和不同氮源的利用情况(图8)可知，以NH₄⁺-N为唯一氮源培养时，菌株在0-12 h进入对数生长期，48 h时有99.80%的NH₄⁺-N被去除。同时检测到8.55 mg/L的NO₃^--N和1.01 mg/L的NO₂^--N以及41.89 mg/L的Cell-N，通过氮平衡计算，有35.84%和35.91%的氮分别被转化为Cell-N和气态氮。以NO₃^--N为唯一氮源时，培养48 h时菌株OD₆₀₀达到1.1，同时，81.54%的NO₃^--N被降解，NO₂^--N的累积量为31.04 mg/L，有44.84%的氮被转化为Cell-N，含量为48.52 mg/L；20.90%的氮被转化为气态氮。以NO₂^--N为唯一氮源时，菌株在36 h内呈现快速生长趋势，NO₂^--N浓度同步降低，48 h去除率达80.00%，有18.99%的氮被转化为气态氮；41.32%的氮被转化为Cell-N，含量为47.84 mg/L。上述不同氮源下菌株的代谢特性证明了硝化反硝化、同化或异化NO₃^--N还原和氨同化途径的存在。

以NH₄⁺-N和NO₃^--N为混合氮源时，12 h NH₄⁺-N去除率为80.86%，NO₃^--N去除率为23.66%。48 h时菌株OD₆₀₀为1.5，NH₄⁺-N和NO₃^--N被完全去除。在NH₄⁺-N和NO₂^--N共存时，12 h NH₄⁺-N去除率达75.14%，NO₂^--N去除率为53.73%。48 h后NH₄⁺-N去除率为100.00%，NO₂^--N去除率为91.97%。综上所述，说明菌株WH-E在混合氮源条件下优先利用NH₄⁺-N进行硝化作用。

2.5 Klebsiellasp. WH-E 脱氮途径的推测

全基因组圈图如图9所示，注释结果表明，菌株WH-E的G+C含量为55.21%，并含有2个质粒。经编码序列预测，共识别出5 283个蛋白编码基因，其编码区总长度达708 664 bp，单个基因平均长度为961.83 bp，编码区域占整个基因组的87.80%。将菌株WH-E测序数据上传至NCBI，获得基因登录号PRJNA1262665。

通过基因组功能分析，在Klebsiella sp. WH-E中鉴定到gdhA、glnA、glnB、nxrAB、norVWR、narGHI、nasBC、nirBD等30余个氮代谢相关基因，并系统阐明了具体的氮代谢途径(图10)。氨同化途径主要通过2条通路实现：(1) 谷氨酸脱氢酶途径，由gdhA基因编码的谷氨酸脱氢酶直接催化α-酮戊二酸和氨生成谷氨酸；(2) 谷氨酰胺合成酶-谷氨酸合成酶途径，由glnA基因编码的谷氨酰胺合成酶催化氨与谷氨酸合成谷氨酰胺，随后该产物在谷氨酸合成酶的催化下重新转化为谷氨酸，实现氨的同化过程。其中，glnA基因的转录活性由glnB、glnL和glnG基因调节，维持氮代谢的平衡^[17]。gltB和gltD基因通过催化谷氨酰胺合成谷氨酸，参与氮素循环和贮存，并转化为α-酮戊二酸，进入三羧酸循环(tricarboxylic acid cycle, TCA)释放能量^[18]。

硝化反硝化途径：在硝化过程中NH₄⁺-N被依次转化为NH₂OH、NO₂^--N和NO₃^--N，其中亚硝酸盐还原步骤由nxrAB基因编码的亚硝酸还原酶催化；在反硝化阶段，NO₃^--N通过由nrtABC基因编码的硝酸盐-亚硝酸盐转运系统进入细胞，并在由narGHIL基因编码的硝酸盐还原酶作用下被还原为NO₂^--N，随后由一氧化氮还原酶(norVWR)基因催化生成一氧化氮(nitric oxide, NO)、N₂O，最后转化为N₂释放。然而，在该菌株基因组中未发现反硝化过程所必需的关键基因nirK/nirS和nosZ基因，这提示菌株WH-E反硝化途径在基因组构成上不同于典型菌株，可能存在新的途径或酶系统参与菌株的氮代谢^[7,11,19]，功能试验证实了其具备将氮氧化物还原为N₂的活性。综上所述，WH-E拥有一条非典型的反硝化途径。

硝酸盐同化与异化途径：通过对硝酸盐还原相关基因的功能注释解析可知，NO₃^--N可经过硝酸盐还原酶/亚硝酸盐氧化还原酶基因(narGHI)或同化性硝酸盐还原酶基因(nasBC)转化为NO₂^--N，并通过亚硝酸盐还原酶基因(nirBD)还原为NH₄⁺-N。narGHI/nirBD基因以及nasA基因分别被认定为是异化硝酸盐还原和同化硝酸盐还原过程的核心功能基因。因此，WH-E可能同时具备上述2条硝酸盐还原通路。

3 讨论

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3.1 Klebsiella sp. WH-E 脱氮特性的影响因素

柠檬酸钠是WH-E菌株的最适碳源，可直接进入TCA供能并合成中间体^[20]，其分解产物有助于维持pH稳定^[21]，相比糖酵解途径代谢的葡萄糖和蔗糖等碳源更有利于菌株脱氮。本研究中14.53的最适C/N比处于HN-AD菌的适宜C/N比10-15范围^[22]，与琼氏不动杆菌(Acinetobacter junii)YB在C/N=15时脱氮性能最优^[23]的报道一致，表明此范围可平衡碳氮供给，避免反硝化不完全或出现资源浪费与菌群抑制现象^[24-25]。菌株WH-E在34.18 ℃时脱氮性能最佳，该温度处于HN-AD菌株最适范围(20-40 ℃)内^[26]，既能维持较高的酶活性，又能避免蛋白质变性^[27]，有利于菌株生长和脱氮性能的发挥。菌株的最适转速为159.59 r/min，此转速可提供适宜的溶氧水平，既能满足好氧代谢需求，又能避免溶氧不足或剪切力过高对菌体造成不利影响^[28]，从而保障高效脱氮。菌株WH-E在初始pH 7.1时表现最优，该值处于HN-AD菌最适pH范围(7.0-7.5)内^[29]，有助于维持代谢活性和酶功能，优化其脱氮性能。为进一步揭示溶氧水平对脱氮产物的调控作用，后续将构建在线DO监测与控制系统，动态解析溶解氧对菌株氮代谢的影响。

3.2 Klebsiellasp. WH-E氮代谢途径的机制分析

结合菌株WH-E在培养过程中的氮素含量变化、气态产物生成情况以及全基因组分析结果，推断菌株WH-E主要具备3种氮代谢途径。在分别以NH₄⁺-N、NO₃^--N、NO₂^--N为氮源的培养条件下，培养48 h后检测到NO₃^--N、NO₂^--N、NH₄⁺-N、N₂、N₂O和Cell-N的累积，说明WH-E能够将NH₄⁺-N转化为NO₂^--N和NO₃^--N，并将NO₃^--N依次转化为NO₂^--N、N₂O和N₂。此外，还可能存在同化或异化NO₃^--N还原途径，将NO₂^--N还原为NH₄⁺-N，随后通过氨同化过程将其固定为生物氮。基因组中含有氨同化相关基因，这表明Klebsiella sp. WH-E能够利用NH₄⁺-N进行同化作用合成蛋白质和核酸，以维持基础代谢和支持细胞生长^[30]。该途径可在养殖废水处理中避免亚硝酸盐的累积。基因组中存在的异化NO₃^--N还原途径的关键酶基因(narGHI和nirBD)和同化还原途径的关键酶基因(nasA)^[31]，表明该菌株可通过同化或异化途径还原NO₃^--N，合成微生物并获取能量^[32]，能够灵活选择代谢途径以提高脱氮效率。菌株WH-E虽然表现出良好的HN-AD能力，但全基因组数据中部分硝化和反硝化关键基因缺失，表明其具有不完全的反硝化途径。这与Pseudomona sp. G16^[33]、食油不动杆菌(Acinetobacter oleivorans)AHP123^[7]和Klebsiella sp. TSH15^[11]的情况类似。由此推测，该菌株可能通过非典型氮转化途径实现脱氮，并且可能有未明确的基因取代了传统硝化-反硝化途径的关键基因，从而形成了独特的氮代谢机制。

3.3 Klebsiellasp. WH-E脱氮优势

研究表明，多数HN-AD菌采用与典型的自养硝化-反硝化路径高度相似的全程硝化反硝化作为代谢通路^[34]。部分HN-AD菌采用直接氨氧化途径和同化/异化硝酸盐还原途径等其他氮代谢途径，这与Halomonas sp. DN3^[9]和施氏假单胞菌(Pseudomonas stutzeri) T13^[29]菌株的情况一致。如表4所示，Klebsiella sp. WH-E比其他HN-AD菌表现出更高的脱氮效率。首先，菌株WH-E具备氨同化以及同步实现硝化和反硝化的功能，无需满足传统脱氮工艺过程中对缺氧环境和复杂时空分区的要求，提高了氮素去除的效率^[38]。其次，该菌可通过同化/异化硝酸盐还原途径来维持能量代谢^[39]，并能根据外界环境因子的改变动态调节硝化作用和氨同化之间的比重，进而增强其生态适应力和生存优势。这凸显了Klebsiella sp. WH-E在高氮负荷废水处理中的应用价值，并为后续的工程化菌剂开发提供了坚实的理论支撑。

4 结论

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本研究从猪场污泥中筛选出一株脱氮性能良好的HN-AD菌Klebsiella sp. WH-E，其最适培养参数为：碳源为柠檬酸钠，温度为34.18 ℃，pH为7.1，C/N为14.53，转速为159.59 r/min。基因组分析证实Klebsiella sp. WH-E具备氨同化途径、硝化反硝化途径和同化/异化硝酸盐还原途径3条主要的氮代谢通路，表现出在畜禽养殖污水脱氮处理中的应用潜力。然而，相关代谢途径尚缺乏关键功能基因，且脱氮性能评估仍局限于纯培养体系。后续可结合基因编辑技术解析非典型氮代谢途径中的关键功能基因，并推动在真实废水体系中的应用验证，深入探究其脱氮机制。

基金

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甘肃省自然科学基金(24JRRA644)
甘肃省科技技术创新引导计划-东西部科技协作专项(25CXNA029)
甘肃农业大学伏羲杰出人才项目(GAUfx-04J03)
甘肃甘味生猪优势特色产业集群科技支撑和技术研发服务项目(GNKJ-2024-46)
甘肃省现代农业产业技术体系建设专项资金(GSARS02)

参考文献

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2026年第66卷第5期

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文章信息

doi: 10.13343/j.cnki.wsxb.20250722

接收时间：2025-09-24
首发时间：2026-05-09
出版时间：2026-05-04

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出版历史

收稿日期：2025-09-24
录用日期：2026-01-26

基金

The Natural Science Foundation of Gansu Province(24JRRA644)

甘肃省自然科学基金(24JRRA644)

The Gansu Science and Technology Innovation Guidance Plan-Eastern-Western Regional Science and Technology Cooperation Initiative(25CXNA029)

甘肃省科技技术创新引导计划-东西部科技协作专项(25CXNA029)

The Fuxi Outstanding Talent Program of Gansu Agricultural University(GAUfx-04J03)

甘肃农业大学伏羲杰出人才项目(GAUfx-04J03)

The Research and Application of Supporting Technologies for Gansu Gansi Pork Advantageous and Characteristic Industry Cluster(GNKJ-2024-46)

甘肃甘味生猪优势特色产业集群科技支撑和技术研发服务项目(GNKJ-2024-46)

The Special Fund for the Construction of the Modern Agricultural Industry Technology System in Gansu Province(GSARS02)

甘肃省现代农业产业技术体系建设专项资金(GSARS02)

作者信息

^1.甘肃农业大学动物科学技术学院，甘肃兰州

^2.甘肃省畜牧技术推广总站，甘肃兰州

^3.平凉市农产品质量安全与检验检测中心，甘肃平凉

^4.临夏回族自治州畜牧技术推广站，甘肃临夏

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2种不同金属材料的力学参数

科 Family	属数 Number of genus	种数 Number of species	占总种数比例 Percentage of total species (%)	属 Genus	种数 Number of species	占总种数比例 Percentage of total species (%)
鹅膏菌科Amanitaceae	2	11	5.26	鹅膏菌属 Amanita	10	4.78
小菇科 Mycenaceae	2	12	5.74	丝盖伞属 Inocybe	5	2.39
多孔菌科 Polyporaceae	8	14	6.70	蜡蘑属 Laccaria	5	2.39
红菇科 Russulaceae	3	23	11.00	小皮伞属 Marasmius	6	2.87
				小菇属 Mycena	11	5.26
				光柄菇属 Pluteus	5	2.39
				红菇属 Russula	17	8.13
				栓菌属 Trametes	5	2.39

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TxT

Test number	Level	NH₄⁺-N removal efficiency/%
1	25	7.0	15	180	71.55
2	25	7.0	20	160	78.62
3	25	6.0	15	160	71.41
4	25	7.0	15	140	70.13
5	25	8.0	15	160	79.59
6	25	7.0	10	160	69.85
7	30	6.0	20	160	81.44
8	30	6.0	15	180	80.86
9	30	7.0	20	180	89.17
10	30	6.0	15	140	79.59
11	30	6.0	10	160	76.55
12	30	8.0	15	180	82.81
13	30	7.0	10	140	79.54
14	30	7.0	15	160	96.87
15	30	7.0	15	160	99.97
16	30	7.0	15	160	94.03
17	30	7.0	15	160	92.88
18	30	7.0	15	160	99.01
19	30	8.0	10	160	84.42
20	30	7.0	20	140	78.43
21	30	8.0	15	140	80.94
22	30	7.0	10	180	80.52
23	30	8.0	20	160	83.71
24	35	7.0	15	180	82.72
25	35	6.0	15	160	80.00
26	35	7.0	15	140	86.61
27	35	8.0	15	160	85.20
28	35	7.0	20	160	80.43
29	35	7.0	10	160	83.94

Test number

Level

NH₄⁺-N removal efficiency/%

T/°C

C/N

Shaking speed/(r/min)

7.0

180

71.55

7.0

160

78.62

6.0

160

71.41

7.0

140

70.13

8.0

160

79.59

7.0

160

69.85

6.0

160

81.44

6.0

180

80.86

7.0

180

89.17

6.0

140

79.59

6.0

160

76.55

8.0

180

82.81

7.0

140

79.54

7.0

160

96.87

7.0

160

99.97

7.0

160

94.03

7.0

160

92.88

7.0

160

99.01

8.0

160

84.42

7.0

140

78.43

8.0

140

80.94

7.0

180

80.52

8.0

160

83.71

7.0

180

82.72

6.0

160

80.00

7.0

140

86.61

8.0

160

85.20

7.0

160

80.43

7.0

160

83.94

Source	Sum of squares	df	Mean square	F-value	P-value	Significant
Model	1 664.76	14	118.91	15.51	<0.000 1	**
A-temperature	277.99	1	277.99	36.25	<0.000 1	**
B-pH	63.88	1	63.88	8.33	0.012 0	*
C-C/N	23.98	1	23.98	3.13	0.098 7	-
D-shaking speed	11.06	1	11.06	1.44	0.249 7	-
AB	2.23	1	2.23	0.29	0.598 2	-
AC	37.69	1	37.69	4.92	0.043 7	*
AD	7.03	1	7.03	0.92	0.354 5	-
BC	7.81	1	7.81	1.02	0.329 9	-
BD	0.53	1	0.53	0.07	0.796 4	-
CD	23.83	1	23.83	3.11	0.099 8	-
A²	736.20	1	736.20	96.01	<0.000 1	**
B²	360.13	1	360.13	46.97	<0.000 1	**
C²	348.39	1	348.39	45.43	<0.000 1	**
D²	405.52	1	405.52	52.89	<0.000 1	**
Residual	107.35	14	7.67	-	-	-
Lack of fit	69.68	10	6.97	0.74	0.682 6	-
Pure error	37.67	4	9.42	-	-	-
Cor total	1 772.11	28	-	-	-	-

Source

Sum of squares

Mean square

F-value

P-value

Significant

Model

1 664.76

118.91

15.51

<0.000 1

A-temperature

277.99

36.25

<0.000 1

B-pH

63.88

8.33

0.012 0

C-C/N

23.98

3.13

0.098 7

D-shaking speed

11.06

1.44

0.249 7

2.23

0.29

0.598 2

37.69

4.92

0.043 7

7.03

0.92

0.354 5

7.81

1.02

0.329 9

0.53

0.07

0.796 4

23.83

3.11

0.099 8

A²

736.20

96.01

<0.000 1

B²

360.13

46.97

<0.000 1

C²

348.39

45.43

<0.000 1

D²

405.52

52.89

<0.000 1

Residual

107.35

7.67

Lack of fit

69.68

6.97

0.74

0.682 6

Pure error

37.67

9.42

Cor total

1 772.11

Nitrogen source	Initial nitrogen concentration/(mg/L)	c(TN)/(mg/L)	Cell-N	Gaseous-N (N₂+N₂O)
NH₄⁺-N	134.07±0.27	10.77±0.17	0.72±0.36	116.87±4.85	64.09±0.21	41.89±1.71	35.84	41.97	35.91
NO₃^--N	2.98±1.08	98.35±3.17	0.22±0.04	108.21±6.45	81.18±0.42	48.52±0.39	44.84	22.62	20.90
NO₂^--N	11.57±2.24	0.00±0.00	91.81±6.28	115.77±0.27	83.45±0.32	47.84±1.73	41.32	21.98	18.99

Nitrogen source

Initial nitrogen concentration/(mg/L)

c(TN)/(mg/L)

Cell-N

Gaseous-N (N₂+N₂O)

NH₄⁺-N

NO₃^--N

NO₂^--N

Initial

Final

c(Cell-N)/(mg/L)

Proportion/%

c(Gaseous-N)/(mg/L)

Proportion/%

NH₄⁺-N

134.07±0.27

10.77±0.17

0.72±0.36

116.87±4.85

64.09±0.21

41.89±1.71

35.84

41.97

35.91

NO₃^--N

2.98±1.08

98.35±3.17

0.22±0.04

108.21±6.45

81.18±0.42

48.52±0.39

44.84

22.62

20.90

NO₂^--N

11.57±2.24

0.00±0.00

91.81±6.28

115.77±0.27

83.45±0.32

47.84±1.73

41.32

21.98

18.99

Bacterial genus name	NH₄⁺-N removal efficiency/%	NO₃^--N removal efficiency/%	NO₂^--N removal efficiency/%	N₂ production	Genes related to nitrogen metabolism	References
Acinetobacteroleivorans AHP123	97.93	-	-	Heterotrophic nitrification: 12.4 mg/L Aerobic denitrification: 18.33 mg/L	nar, amt, glnK, gltB, gdhA, gam, glnA, ncd2, ncd1, nirB, nasA	[7]
Klebsiella sp. TSH15	92.55	91.43	-	-	amt, glnA, gdhA, nirB, narG, napA, nasA, nirK/S, norB, nosZ	[11]
Candida boidinii L21	96.00	98.00	-	-	-	[12]
Pseudomonas aeruginosa WS-03	95.81	94.00	100.00	-	narB, glnA, gdhA, nirB, nirA, napA, gltB	[19]
Acinetobacter ZQ‑A1	84.84	87.13	92.63	-	-	[35]
Pseudomonas sp. Y-5	97.26	99.34	95.81	-	-	[36]
Pseudomonas mendocina TJPU04	98.00	93.00	100.00	-	-	[37]
Klebsiella sp. WH-E	99.80	81.54	80.00	With NH₄⁺-N as the sole nitrogen source: 35.91% With NO₂^--N as the sole nitrogen source: 20.90% With NO₃^--N as the sole nitrogen source: 18.99%	glnB, glnL, glnG, gdhA, norVWR, narGHI, nasBC, nirBD

Bacterial genus name

NH₄⁺-N

removal efficiency/%

NO₃^--N

removal efficiency/%

NO₂^--N

removal efficiency/%

N₂ production

Genes related to nitrogen metabolism

References

Acinetobacteroleivorans AHP123

97.93

Heterotrophic nitrification: 12.4 mg/L

Aerobic denitrification: 18.33 mg/L

nar, amt, glnK, gltB, gdhA, gam, glnA, ncd2, ncd1, nirB, nasA

[7]

Klebsiella sp. TSH15

92.55

91.43

amt, glnA, gdhA, nirB, narG, napA, nasA, nirK/S, norB, nosZ

[11]

Candida boidinii L21

96.00

98.00

[12]

Pseudomonas

aeruginosa WS-03

95.81

94.00

100.00

narB, glnA, gdhA, nirB, nirA, napA, gltB

[19]

Acinetobacter ZQ‑A1

84.84

87.13

92.63

[35]

Pseudomonas sp. Y-5

97.26

99.34

95.81

[36]

Pseudomonas

mendocina TJPU04

98.00

93.00

100.00

[37]

Klebsiella sp. WH-E

99.80

81.54

80.00

With NH₄⁺-N as the sole nitrogen source: 35.91%

With NO₂^--N as the sole nitrogen source: 20.90%

With NO₃^--N as the sole nitrogen source: 18.99%

glnB, glnL, glnG, gdhA, norVWR, narGHI, nasBC, nirBD