Identification of undesirable white-colony-forming yeasts appeared on the surface of Japanese kimchi

Figures & data

Figure 1. Changes in pH (A) and viable cell counts of LAB (B) and yeast (C) in kimchi A (circles), kimchi B (triangles), and kimchi C (squares). Kimchi was fermented at 4 °C (open symbols) or 15 °C (filled symbols) for 28 days. Viable cell counts of LAB and yeast were determined with GYP and YPD agar media, respectively.

Figure 2. PCR-DGGE analysis of yeast communities in three types of kimchi fermented at 4 or 15 °C. Bands labeled with letters were excised and their DNA sequences were determined for identification of yeasts (see Table 1). The numbers above each lane represent the fermentation period (days).

Table 1. Sequence analysis of 26S rRNA genes recovered from DGGE bands.

Band	Length (base)	Most closely related organism in GenBank database	Accession No.	% Similarity
a	186	Candida sake	EF017662	100
b	172	Debaryomyces hansenii	KY511974	99
c	193	Geotrichum fragrans	U40119	100
d	109	Candida picinguabensis	AB437381	100
e	200	Debaryomyces hansenii	KY511974	100
f	175	Kazachstania exigua	JQ808008	100
g	189	Trichosporon sp.	LC034289	99
h	177	Debaryomyces hansenii	KY107560	100
i	190	Trichosporon shinodae	AB180201	100
j	189	Brassica oleracea var. capitata	KT377455	98
k	191	Brassica oleracea var. capitata	KT377455	98
l	196	Trichosporon dermatis	KT981961	100
m	201	Brassica oleracea var. capitata	KT377455	97
n	162	Candida sake	U45728	100
o	175	Kazachstania exigua	JQ808008	100
p	202	Debaryomyces hansenii	KY511974	100
q	171	Geotrichum fragrans	U40119	100
r	171	Geotrichum fragrans	U40119	100
s	192	Debaryomyces hansenii	KY512470	100
t	193	Debaryomyces hansenii	KY511967	100
u	178	Pichia fermentans	KJ413165	100
v	178	Pichia fermentans	KJ413165	100
w	175	Kazachstania exigua	JQ808008	100

Figure 3. PCR-DGGE analysis of yeast communities in three types of kimchi after white-colony formation. Kimchi was fermented at 4, 10, 15, or 25 °C until white colonies appeared on the surface. Note that white-colony formation at 4 °C was detected only in kimchi C. Bands labeled with letters were excised, and their DNA sequences were determined for identification of yeasts (see text). Numbers above each lane represent fermentation temperature (°C).

Table 2. Yeast strains isolated directly from white colonies formed on kimchi surface.

Fermentation temperature (°C)	Most closely related organism in GenBank database	Number of strains
4^a	Candida sake	6
10	Kazachstania pseudohumilis	7
	Candida sake	2
	Kazachstania exigua	2
15	Kazachstania exigua	7
	Kazachstania pseudohumilis	1
25	Kazachstania exigua	11
	Pichia fermentans	6
	Kazachstania pseudohumilis	1

^a Of three kimchi products, white film formation at 4 °C occurred only in kimchi C after a prolonged fermentation period of 35 days.

Figure 4. White-colony formation on the surface of kimchi A inoculated with yeast strains. Freshly prepared kimchi A was heat-treated and inoculated with 10² CFU g⁻¹ of Kazachstania exigua (B and E) or K. pseudohumilis (C and F). Control kimchi without yeast inoculation is also shown (A and D). The kimchi was incubated at 15 °C for 21 days (panels A–C) or at 4 °C for 51 days (D–F). Two strains were inoculated individually for each yeast species, but essentially the same results were obtained. White arrows indicate the yeast colonies.

Table 3. The growth rates (μ) of yeast strains at 4 and 15 °C in YPD medium.^a

Temp. (°C)	Acidulants^b(%)	K. exigua	K. pseudohumilis	C. sake	P. fermentans	D. hansenii	T. moniliforme	C. picinguabensis
15	0	0.632	0.590	0.479	0.475	0.230	0.105	0.375
	0.2	0.530	0.499	0.420	0.372	0.324	0.103	0.229
	0.4	0.492	0.449	0.202	0.372	0.076	0	0
4	0	0	0	0.157	0.159	0.148	0.098	0

^a Average data obtained from two different strains are shown. For T. moniliforme and C. picinguabensis, only one strain was used.

^b A mixture of acidulants was added to the medium at final concentrations of 0.2 to 0.4% (w/v).