Characterization of flux distribution and general physiology of S. pyogenes in continuous cultures

Selected Cell

Cell:

Value:

Metadata Organism_sample Data_ExtracellularMetabolites Results_CalculatedFluxes Instrument_chemostat Instrument_measurements_envPara MediumComposition

Metadata	Values
Asset Title	Characterization of flux distribution and general physiology of S. pyogenes in continuous cultures
Uploader	Martijn Bekker
Uploader SEEK ID	53
Project	SysmoLab

ASSAY
Assay SEEK ID	109
Assay Title	Physiological characterization of Lactic acid bacteria grown in C-limited continuous cultures
Assay_type	catabolic flux distribution
Technology_type	HPLC
Description	S. pyogenes was grown in C-limited cultures at pH 6.7 and 7.5 and at a growth rate of 0.05 and 0.15
Experimentalist	Martijn Bekker, Tomas Fiedler
Date	2010-08-01T0:0:0+0000
SOP	2


Experimental_conditions
Item	temperature	Growth_Medium	Growth_Medium
Compound (if concentration)		Glucose	Acetate
Unit	ºC	mM	mM
Start_value (optional)		65	12
End_value (optional)	37
Comments		standard medium with additional glucose	standard medium with additional acetate
Culture Growth	Chemostat
FACTORS_STUDIED
Item	pH	dilution_rate
Compound (if concentration)
Unit		1/h
Start_value (optional)	6.5	0.05
End_value (optional)	7.5	0.15
SD (optional)
Comments		Growth rate dependent flux distribution

Sample							Treatment(s)		Culture_start_date	Date_at_sampling	Age_at_sampling_hours
	Organism	NCBI_ID	Strain	Genotype	Phenotype	organism/cell part	pH	Dilution_rate	date	date	age
1	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	6.5	0.05
2	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	6.5	0.15
3	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	7.5	0.05
4	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	7.5	0.15
5	Streptococcus_pyogenes	1314	M49	Δldh		whole cell	6.5	0.05
6	Streptococcus_pyogenes	1314	M49	Δldh		whole cell	6.5	0.15
7	Streptococcus_pyogenes	1314	M49	Δldh		whole cell	7.5	0.05
8	Streptococcus_pyogenes	1314	M49	Δldh		whole cell	7.5	0.15
9	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	6.5	0.05
10	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	7.5	0.05
11	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	7.5	0.05
11a	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	7.5	0.15
12	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	7.5	0.05
13	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	7.5	0.15
13a	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	7.5	0.05
14	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	6.5	0.15
15	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	6.5	0.05
16	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	6.5	0.05
17	Streptococcus_pyogenes	1314	M49	wild-type		whole cell	6.5	0.15

Metabolite	Sample	Glucose	Ethanol	Acetate	Lactate	Formiate	Succinate	Pyruvate	Acetoin	2,3 Butanediol	CO2	O2	Aspartic acid	Serine	Glutamate	Glycine	Histidine	NH3	Arginine	Threonine	Alanine	Proline	Cysteine	Tyrosine	Valine	Methionine	Ornithine	Lysine	Isoluecine	Leucine	Phenylalanine
Metabolite ID		CHEBI:17234	CHEBI:16236	CHEBI:30089	CHEBI:24996	CHEBI:15740	CHEBI:30779	CHEBI:15361	CHEBI:15688	CHEBI:62064	CHEBI:16526	CHEBI:15379	CHEBI:22660	CHEBI:17822		CHEBI:15428	CHEBI:27570	CHEBI:16134	CHEBI:29016	CHEBI:26986	CHEBI:16449	CHEBI:26271	CHEBI:15356	CHEBI:18186	CHEBI:27266	CHEBI:16811	CHEBI:18257	CHEBI:25094	CHEBI:24898	CHEBI:25017	CHEBI:28044
Unit		mM	mM	mM	mM	mM	mM	mM	mM	mM	%	%	mM	mM	mM	mM	mM	mM	mM	mM	mM	mM	mM	mM	mM	mM	μM	mM	mM	mM	mM
	1	0	19.3	35.1	70	35.4	0	1.5	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	2	0	0	20.3	99.7	8.1	0	1.4	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	3	0	8.3	22.8	83.8	13.6	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	4	0	11	20.2	56.2	20.7	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	5	0	44.1	40.3	1.8	65.4	0	8.2	3.2	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	6	0	55.7	42.1	2.3	74.6	0	19.5	3	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	7	0	61.1	40.8	1.7	71.5	0	15.4	2.6	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	8	0	55.3	39.6	0.8	69	0	15	2.5	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	9	0	28.5996	26.5911	33.2897	27.1529	2.7314	0.2222	9.0482	0
	10	0	32.5404	58.4248	12.1797	71.7257	4.2334	0.2573	2.9294	0
	11	0	16.9109	46.1068	34.7783	45.5907	0	0	1.7379	0
	11a	0	4.5031	25.3673	65.3299	11.4123	0	0	1.7817	0
	12	0	0	0	0	0	0	0	0	0
	13	4.8704	0	16.1389	85.2263	0	0	0	1.0621	0
	13a	0	12.0724	35.0679	66.9688	26.5225	0	0	1.6499	0
	14	0	0	15.1085	96.3673	0	0	0	1.178	0
	15	0	6.8169	32.5884	69.796	17.6214	0.1566	0.806	0	0
	16	0	0	35.4929	75.7861	0	0.6109	0.4942	0	0
	17	0	11.4256	44.4907	68.0624	38.2789	0.3511	0	0	0

		Metabolite
Sample	Dilution rate	Glucose	Ethanol	Acetate	Lactate	Formate	Succinate	Pyruvate	Acetoin	2,3-butanediol	CO2	O2	Aspartic acid	Serine	Glutamate	Glycine	Histidine	NH3	Arginine	Threonine	Alanine	Proline	Cysteine	Tyrosine	Valine	Methionine	Ornithine	Lysine	Isoluecine	Leucine	Phenylalanine	Yield glucose	Yield glucose	ATP (calculated)	Lactate (mol/mol glucose)	Formate (mol/mol glucose)	EtOH (mol/mol glucose)	Acetate (mol/mol glucose)	Acetoin (mol/mol glucose)	EtOH+ Acetate (mol/mol glucose)	Carbon balance	Redox balance	PDH flux	Relative PDH flux
		CHEBI:17234	CHEBI:16236	CHEBI:30089	CHEBI:24996	CHEBI:15740	CHEBI:30779	CHEBI:15361	CHEBI:15688	CHEBI:62064	CHEBI:16526	CHEBI:15379	CHEBI:22660	CHEBI:17822		CHEBI:15428	CHEBI:27570	CHEBI:16134	CHEBI:29016	CHEBI:26986	CHEBI:16449	CHEBI:26271	CHEBI:15356	CHEBI:18186	CHEBI:27266	CHEBI:16811	CHEBI:18257	CHEBI:25094	CHEBI:24898	CHEBI:25017	CHEBI:28044
Unit	/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	mmol/gram/hour	g/mol	g/g	mmol/gram/hour	mol/mol	mol/mol	mol/mol	mol/mol	mol/mol	mol/mol	%	mol NADH produced-mol NADH consumed	mmol/gram/hour	percentage
1	0.05	-2.7083333333333335	0.8041666666666668	0.9625	2.916666666666667	1.475	0	0.06250000000000001	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	13.076923076923077	0.07264957264957264	5.708333333333334	1.0769230769230769	0.5446153846153846	0.29692307692307696	0.35538461538461535	0	-0.6523076923076924	0.8761538461538462	0.5125	0.29166666666666674	0.16509433962264153
2	0.15	-13.928571428571429	0	1.778571428571429	21.364285714285717	1.7357142857142855	0	0.3	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	12.102564102564102	0.06723646723646723	25.221428571428575	1.533846153846154	0.1246153846153846	0	0.12769230769230772	0	-0.12769230769230772	0.8415384615384617	2.1214285714285723	0.04285714285714337	0.02409638554216896
3	0.05	-5	0.6384615384615385	0.8307692307692308	6.446153846153846	1.0461538461538462	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	11.384615384615385	0.06324786324786325	8.746153846153847	1.2892307692307692	0.20923076923076925	0.12769230769230772	0.16615384615384615	0	-0.29384615384615387	0.7915384615384615	0.6153846153846154	0.42307692307692313	0.2879581151832461
4	0.15	-9.75	1.65	1.2299999999999998	8.43	3.105	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	10.76923076923077	0.05982905982905983	12.54	0.8646153846153846	0.31846153846153846	0.1692307692307692	0.12615384615384612	0	-0.29538461538461536	0.58	-0.6450000000000002	-0.2250000000000001	-0.07812500000000003
5	0.05	-3.25	2.205	1.415	0.09000000000000001	3.2700000000000005	0	0.41	0.16000000000000003	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	11.641025641025642	0.06467236467236467	5.855	0.027692307692307697	1.0061538461538464	0.6784615384615384	0.43538461538461537	0.04923076923076924	-1.113846153846154	0.6830769230769231	0.28999999999999965	0.34999999999999964	0.09668508287292808
6	0.15	-7.8	6.684	3.6119999999999997	0.27599999999999997	8.952	0	2.34	0.36	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	15.076923076923077	0.08376068376068375	17.244	0.03538461538461538	1.1476923076923078	0.856923076923077	0.46307692307692305	0.046153846153846156	-1.3199999999999998	0.8738461538461538	1.3319999999999987	1.3439999999999994	0.1305361305361305
7	0.05	-5	4.7	2.215384615384615	0.13076923076923078	5.5	0	1.1846153846153846	0.2	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	8.461538461538462	0.04700854700854701	10.846153846153845	0.026153846153846156	1.1	0.9400000000000001	0.44307692307692303	0.04	-1.383076923076923	0.8630769230769232	0.5153846153846152	1.4153846153846157	0.20467185761957735
8	0.15	-9.285714285714285	7.8999999999999995	3.942857142857142	0.11428571428571428	9.857142857142856	0	2.142857142857143	0.35714285714285715	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	12.461538461538463	0.06923076923076923	18.757142857142856	0.01230769230769231	1.0615384615384615	0.8507692307692308	0.42461538461538456	0.038461538461538464	-1.2753846153846153	0.7976923076923076	0.8857142857142856	1.9857142857142858	0.16767189384800968
9	0.05	-4.887218045112782	2.150345864661654	1.0970751879699248	2.5029849624060154	2.0415714285714284	0.20536842105263156	0.016706766917293236	0.6803157894736841	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	11.666666666666666	0.06481481481481481	8.635556390977444	0.5121492307692309	0.41773692307692306	0.4399938461538461	0.22447846153846154	0.1392030769230769	-0.6644723076923077	0.729223076923077	1.5299172932330825	1.2058496240601504	0.3713253084576078
10	0.05	-7.222222222222222	3.6155999999999997	5.158311111111111	1.3533000000000002	7.969522222222222	0.4703777777777778	0.028588888888888886	0.3254888888888889	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	27.692307692307693	0.15384615384615385	16.905844444444448	0.18738000000000002	1.1034723076923076	0.5006215384615385	0.7142276923076922	0.04506769230769231	-1.214849230769231	0.7481615384615385	3.026666666666667	0.804388888888889	0.0916796259618161
11	0.05	-5.613636363636301	1.460486818181802	2.94558727272724	3.003580454545421	3.9373786363635928	0	0	0.15009136363636197	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	11.743589743589745	0.06524216524216525	10.655424545454427	0.5350507692307692	0.7013953846153846	0.2601676923076923	0.5247200000000001	0.026736923076923078	-0.7848876923076924	0.686706153846154	2.2539786363636116	0.4686954545454496	0.10637484637684573
11a	0.15	-11.470588235294118	0.7946647058823529	2.358935294117647	11.52880588235294	2.013935294117647	0	0	0.3144176470588236	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	13.102564102564104	0.0727920227920228	17.67017647058823	1.0050753846153844	0.17557384615384616	0.06927846153846154	0.20565076923076922	0.027410769230769235	-0.27492923076923076	0.6674130769230767	3.332770588235294	1.1396647058823528	0.36138530754767656
12	0.05	-7.386363636363637	0	-1.3636363636363638	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	6.717948717948718	0.037321937321937323	-2.7272727272727275	0	0	0	-0.18461538461538463	0	0.18461538461538463	-0.09230769230769231	-2.7272727272727275	-1.3636363636363638	1
13	0.15	-12.355397260273973	0	0.8504589041095889	17.512253424657533	0	0	0	0.21823972602739727	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	11.641025641025642	0.06467236467236467	19.649650684931505	1.4173767994465287	0	0	0.06883298741385273	0.017663513477555147	-0.06883298741385273	0.7607684069077459	2.137397260273972	0.8504589041095889	1
13a	0.05	-4.126984126984206	0.766501587301602	1.4646285714285996	4.251987301587382	1.6839682539682863	0	0	0.10475555555555754	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	10.598290598290596	0.058879392212725534	8.157257142857299	1.0302892307692306	0.4080384615384616	0.18572923076923079	0.3548907692307693	0.02538307692307692	-0.5406200000000001	0.8108376923076923	1.454800000000028	0.5471619047619154	0.24523979590384834
14	0.15	-18.055555555555554	0	0.863472222222222	26.768694444444442	0	0	0	0.3272222222222222	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	8.307692307692308	0.046153846153846156	29.15008333333333	1.4825738461538462	0	0	0.04782307692307691	0.018123076923076922	-0.04782307692307691	0.7833215384615384	2.381388888888888	0.863472222222222	1
15	0.05	-4.421768707482879	0.46373469387753913	1.4005714285713924	4.748027210884232	1.1987346938775203	0.01065306122448952	0.0548299319727877	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	11.299145299145296	0.06277302943969609	8.089040816326325	1.0737846153846158	0.2710984615384616	0.10487538461538465	0.3167446153846154	0	-0.42162000000000005	0.7539023076923078	1.657238095238052	0.6655714285714112	0.35700758612385186
16	0.05	-4.744525547445547	0	1.7148102189782075	5.531832116788662	0	0.04459124087591516	0.036072992700732144	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	9.726495726495726	0.0540360873694207	9.086708029197638	1.1659400000000002	0	0	0.3614292307692308	0	-0.3614292307692308	0.7674861538461539	3.465693430657147	1.7148102189782075	1
17	0.15	-14.828897338402546	2.6066007604561863	7.4123269961974705	15.527543726235221	8.732828897338111	0.08009885931558669	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	7.076923076923076	0.03931623931623931	33.11899619771752	1.047113846153846	0.5889061538461539	0.17577846153846152	0.49985692307692303	0	-0.6756353846153845	0.8613746153846152	6.09182509505683	1.2860988593155458	0.12836691615641568

	Sample	Age	Date	Chemostat	Medium flow	Volume	Temperature	pH Chemostat	Dilution rate	%O2 input	Airflow	N2 flow	Total flow	Comments
Unit	1	hours	Date	name	ml/hour	ml	Celcius	1	1/hour	volume%	ml/min	ml/min	ml/min	1
	1			Tomas1		750	37	6.5	0.05	0	0	500	100	wild-type
	2			Tomas1		750	37	6.5	0.15	0	0	500	100	wild-type
	3			Tomas1		750	37	7.5	0.05	0	0	500	100	wild-type
	4			Tomas1		750	37	7.5	0.15	0	0	500	100	wild-type
	5			Tomas1		750	37	6.5	0.05	0	0	500	100	Δldh
	6			Tomas1		750	37	6.5	0.15	0	0	500	100	Δldh
	7			Tomas1		750	37	7.5	0.05	0	0	500	100	Δldh
	8			Tomas1		750	37	7.5	0.15	0	0	500	100	Δldh
	9	144	2009-11-09T0:0:0+0000	Sysmo1	50	1000	37	6.5	0.05	0	0	100	100	wild-type
	10	144	2009-11-13T0:0:0+0000	Sysmo1	50	1000	37	7.5	0.05	0	0	100	100	wild-type, way to high OD
	11	144	25-01-2010	Sysmo3	50	1000	37	7.5	0.05	0	0	100	100	wild-type
	11a	168	26-01-2010	Sysmo3	75	500	37	7.5	0.15	0	0	100	100	wild-type
	12	96	2010-03-04T0:0:0+0000	SYSMO4	50	1000	37	7.5	0.05	0	0	100	100	wild-type
	13	96	2010-03-05T0:0:0+0000	SYSMO4	150	1000	37	7.5	0.15	0	0	100	100	wild-type
	13a	96	2010-03-06T0:0:0+0000	SYSMO3	50	1000	37	7.5	0.05	0	0	100	100	wild-type
	14	96	2010-03-09T0:0:0+0000	SYSMO4	150	1000	37	6.5	0.15	0	0	100	100	wild-type
	15	96	26-03-2010	SYSMO3	50	1000	37	6.5	0.05	0	0	100	100	wild-type
	16	96	26-03-2010	SYSMO4	50	1000	37	6.5	0.05	0	0	100	100	wild-type
	17	168	29-03-2010	SYSMO4	150	1000	37	6.5	0.15	0	0	100	100	wild-type

Identifier	Sample	CO2	O2	CO2	pH	RPM	OD	standard deviation OD	Dryweight correction calculated	Dry weight	Dry weight	standard deviation dryweight
Unit		volume%	volume%	volume%	1	rpm	AU	AU		gram/liter	gram/liter	gram/liter
Description		Inputgas	Outputgas	Outputgas		stirring rate	Optical density (600nm)	Optical density (600nm)		Calculated	Dryweight measured as described in SOP X
	1					400	2.55		0.47058823529411764	0.85	1.2
	2					400	2.36		0.2966101694915254	0.7866666666666666	0.7
	3					400	2.22		0.2927927927927928	0.7400000000000001	0.65
	4					400	2.1		0.47619047619047616	0.7000000000000001	1
	5					400	2.27		0.44052863436123346	0.7566666666666667	1
	6					400	2.94		0.4251700680272109	0.98	1.25
	7					400	1.65		0.393939393939394	0.5499999999999999	0.65
	8					400	2.43		0.43209876543209874	0.81	1.05
	9	0.01	ND	ND	6.3	400	2.275	0.10606601717796739	0.2923076923076923	0.7583333333333333	0.665	0.24748737341529123
	10	0.01	ND	ND	7.41	400	5.4	0.14142135623729543	0.08333333333333333	1.8	0.45	0.08485281374238561
	11	0.01	ND	ND	7.21	400	2.29	0.1697056274847673	0.25281544472535333	0.7633333333333333	0.5789473684210591	0.029772917102586262
	11a	0.01	ND	ND	7.44	400	2.555	0.10606601717799251	0.33268101761252444	0.8516666666666667	0.85	0.0742462120246034
	12	0.01	ND	ND	7.51	400	1.31	0.15556349186103902	0.33587786259541985	0.4366666666666667	0.44	0.0742462120246034
	13	0.01	ND	ND	7.32	100	2.27	0.09899494936611793	0.32158590308370044	0.7566666666666667	0.73
	13a	0.01	ND	ND	ND	100	2.0666666666666664	0.09073771725877823	0.38104838709676697	0.6888888888888888	0.787499999999985	0.18031222920258855
	14	0.01	ND	ND	6.44	100	1.62	0.3676955262170038	0.3333333333333333	0.54	0.54
	15	0.01	ND	ND	6.46	100	2.203333333333333	0.15567059238448272		0.7344444444444443	0.735000000000019	0.07778174593052666
	16	0.01	ND	ND	6.57	100	1.8966666666666667	0.15044378795195365		0.6322222222222222	0.6849999999999579	0.014142135623761574
	17	0.01	ND	ND	6.69	100	1.38	0.20223748416156723		0.45999999999999996	0.657500000000022	0.003535533905865814

Identifier	Sample	Glucose	Ethanol	Acetate	Lactate	Formiate	Succinate	Pyruvate	Acetoin	2,3 Butanediol	%O2	CO2	Aspartic acid	Serine	Glutamate	Glycine	Histidine	NH3	Arginine	Threonine	Alanine	Proline	Cysteine	Tyrosine	Valine	Methionine	Ornithine	Lysine	Isoluecine	Leucine	Phenylalanine
Unit		mM	mM	mM	mM	mM	mM	mM	mM	mM	%	%	mM	mM	mM	mM	mM	mM	mM	mM	mM	mM	mM	mM	mM	mM	μM	mM	mM	mM	mM
	1	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	2	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	3	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	4	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	5	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	6	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	7	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	8	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	9	65	0	12	0	0	0	0	0	0
	10	65	0	12	0	0	0	0	0	0
	11	65	0	12	0	0	0	0	0	0
	11a	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	12	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	13	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	13a	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	14	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	15	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	16	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	17	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	0	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	0	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	0	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	0	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	0	65	0	12	0	0	0	0	0	0			0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0