Recovering traditional raw-milk Tetilla cheese flavour and sensory attributes by using Kocuria varians and Yarrowia lipolytica adjunct cultures.

The rationale of the present study was to evaluate the potential of microbial adjunct cultures including Kocuria varians and/or Yarrowia lipolytica strains in the recovery of the typical sensory profile of traditional (raw-milk) Tetilla cheese. Four batches of Tetilla cheese, a short ripened cows' milk cheese produced in Galicia (NW Spain), were made in duplicate from pasteurized milk inoculated with different microbial cultures. A control batch was manufactured by adding a mesophilic commercial D-starter only. The other three batches were made with the same starter after a cheese-milk pre-ripening step carried out with (i) an adjunct culture of K. varians, (ii) an adjunct culture of Y. lipolytica, or (iii) a combination of both adjunct cultures. The highest pH and water activity values, associated with softer textures were determined in the cheeses manufactured with the Y. lipolytica adjunct after 21days of ripening. The contents of the volatile compounds 3-methylbutanol, dimethyl disulfide and dimethyl trisulfide were higher in the cheeses made with only the K. varians adjunct than in the cheeses made with the only yeast adjunct and in the control cheeses. The contents of hexanoic and octanoic acids were highest in the cheeses made with the Y. lipolytica adjunct, and levels of ethyl hexanoate, ethyl octanoate and ethyl decanoate were higher in the cheeses made with only the yeast adjunct than in the other batches of cheese. The cheeses manufactured with both adjunct cultures were awarded the highest scores for flavour and overall sensory parameters (considering the standards of the traditional product) and were considered very similar to 'good quality' artisanal raw-milk cheeses. We conclude that use of selected Micrococcaceae and Y. lipolytica strains as adjunct cultures would differentiate the sensory properties and contribute to the quality and typicality of the short-ripened rennet-curd Galician Tetilla and Arzúa-Ulloa cheeses.

Bacteriocin production by Streptococcus thermophilus in complex growth media.

OBJECTIVE: To test if the production of bacteriocins by Streptococcus thermophilus is influenced when grown in various complex media commonly used for the culturing of lactic acid bacteria. RESULTS: Forty-one strains of S. thermophilus were screened for the production of bacteriocins in tryptone/yeast extract/lactose (TYL), M17-lactose (M17L), M17-glucose (M17G) and MRS media. Two strains, ST144 and ST145, were identified as novel bacteriocin producers, with constitutive production observed only in M17G. Strains ST110, ST114 and ST134 constitutively produced bacteriocins in all growth media but ST114 required growth in MRS for its antimicrobial activity to persist in a 24 h culture. The addition of a synthetic quorum sensing peptide (BlpC) induced bacteriocin production by ST106 in all media tested; and by ST118 in TYL and M17L. Strain ST109, which constitutively produced a bacteriocin in TYL and M17 broths, required BlpC induction when grown in MRS. Real-time PCR analysis showed that the natural expression of blpC in ST109 was lower when grown in MRS, suggesting that something in medium interfered with the blp quorum sensing system. CONCLUSION: As the choice of growth medium influences both bacteriocin production and peptide stability, several types of production media should be tested when screening for novel bacteriocin-producing strains of S. thermophilus.

Characterization of yeasts isolated from artisanal short-ripened cows' cheeses produced in Galicia (NW Spain).

A total of 143 presumptive yeast isolates were obtained from the predominant microflora of 21 short-ripened starter-free raw cow's milk cheeses made in Galicia (NW Spain), and the following 68 isolates were identified by both genotyping and sequencing methods: Yarrowia lipolytica (21 isolates), Kluyveromyces lactis (18), Debaryomyces hansenii (11), Pichia guilliermondii (11), Pichia fermentans (4) and Saccharomyces cerevisiae (3). Of these, Y. lipolytica and K. lactis displayed the strongest extracellular proteolytic activity on skim milk agar, and none of the D. hansenii isolates showed any activity on this medium. Y. lipolytica also displayed the highest lipolytic activity on Tween 80 and on tributyrin. This species, which was characterized by production of butanoic acid, free fatty acid esters and sulfur compounds in pasteurized whole milk, was responsible for rancid and cheesy flavors. K. lactis mainly produced acetaldehyde, ethanol, branched chain aldehydes and alcohols, and acetic acid esters, which were responsible for alcoholic, fruity and acetic notes. The volatile profiles of D. hansenii were rather limited and characterized by high levels of methyl ketones. Most of the yeast isolates were described as tryptamine producers, although low concentrations of histamine were produced by five Y. lipolytica and two P. fermentans isolates. We conclude that selected Y. lipolytica strains could be used as adjunct cultures in the manufacture of Arzúa-Ulloa and Tetilla cheeses, and selected K. lactis strains could be used as co-starters in the manufacture of acid curd Cebreiro cheese, thus contributing to the sensory quality and typicality of the cheeses.

Chemical and biochemical study of industrially produced San Simón da Costa smoked semi-hard cow's milk cheeses: Effects of storage under vacuum and different modified atmospheres.

Two batches of smoked, semi-hard (ripened for 45 d) San Simón da Costa cow's milk cheeses with Protected Designation of Origin were used to investigate the chemical, biochemical, and sensorial parameters that may be affected by modified-atmosphere packaging. Cheeses were packaged for 45 d as follows: vacuum packaging, packaging in 100% N(2), packaging in a gas mixture of 20% CO(2)/80% N(2), and packaging in a gas mixture of 50% CO(2)/50% N(2). The San Simón da Costa cheeses were characterized by high contents of lactic, oxalic, and citric organic acids. The main free amino acids found were isoleucine, phenylalanine, serine, valine, lysine, and glutamic acid, and the most abundant volatile compounds included ethanol, diacetyl, 2-butanol, isopropyl alcohol, furfural, acetaldehyde, 2-butanone, acetone, and 2-methylfuran. Modified atmospheres appeared to alter the ripening processes by affecting lipolysis, as indicated by the lower concentrations of butyric and propionic acids compared with control cheeses. In addition, modified-atmosphere packaging altered the proteolysis processes, yielding higher amounts of branched-chain alcohols. The results revealed that storage under modified atmosphere contributes to the accumulation of several compounds probably derived from smoke, including aldehydes such as 2-furancarboxaldehyde (furfural), alcohols such as 2-methoxyphenol (guaiacol), ketones such as 2-cyclopenten-1-one, and esters such as methyl furancarboxylate, which were negatively correlated with flavor. Vacuum packaging was the most useful technique in terms of preserving the sensory quality of San Simón da Costa Protected Designation of Origin cheeses. Considering the current demands for packaged portions of food at the distribution and retail levels and the potential health risks associated with some smoke-derived compounds usually present in some smoked foods, the results obtained in this study may be of special interest to the cheese industry.

Antibiotic resistance in lactic acid bacteria and Micrococcaceae/Staphylococcaceae isolates from artisanal raw milk cheeses, and potential implications on cheese making.

Antibiotic susceptibility against 19 antimicrobial agents was evaluated in isolates of the genera Lactococcus (46 isolates), Leuconostoc (22), Lactobacillus (19), Staphylococcus (8), Enterococcus (7), and Microccoccus/Kocuria (5) obtained from the predominant microflora of nonrecent and recent types of artisanal raw cow's milk cheeses. Beta-lactams showed broad activity against all genera, although leuconostocs and lactobacilli were highly resistant to oxacillin (80% to 95.5%). Resistance to aminoglycosides was frequent for lactococci and enterococci (particularly for streptomycin), whereas lower rates of resistance were detected for lactobacilli and leuconostocs. Technologically interesting traits for the food industry were distributed among isolates that showed different degrees of resistance to common antibiotics. However, isolates showing resistance to less than 2 antibiotics were mainly those with properties of greatest technological interest (acidifying activity, proteolytic/lipolytic activities, or diacetyl production).

Colonization antigens of enterotoxigenic Escherichia coli strains isolated from piglets in Spain.

Eighty-eight enterotoxigenic E.coli strains isolated from 69 pigs with enteric infections (diarrhoea or oedema disease) were investigated for the presence of F4 (K88), F5 (K99), F6 (987P) and F41 colonization antigens. The commonest colonization antigen was F6 (987P), which was detected in ETEC strains from 31.9% pigs, followed by F5 (K99) 11.6%, F4 (K88) 10.1% and F41 8.7%. Presence of F6 (987P) and F5 (K99) fimbriae was statistically associated (0.025 > p < 0.005) with diarrhoeic piglets younger than 15 days. F4 (K88) colonization antigen was only expressed by ETEC isolated from piglets older than 15 days. 90.5% of ETEC isolated from 90.0% of piglets younger than 15 days expressed F5 (K99), F6 (987P) or F41 antigens, whereas only 31.3% ETEC isolated from piglets older than 15 days were positive for F4 (K88), F5 (K99), F6 (987P) or F41 antigens (p < 0.001). None of the ETEC pigs with oedema disease produced any of the four colonization antigens. ETEC bearing colonization antigens were associated with particular serogroups and toxic phenotypes, whereas 4P- ETEC strains showed diverse phenotypic characteristics.

Development and evaluation of an ELISA to detect Escherichia coli K88 (F4) fimbrial antibody levels.

An enzyme-linked immunosorbent assay (ELISA) to determine IgG antibody levels against K88 (F4) fimbrial antigen from porcine enterotoxigenic Escherichia coli (ETEC) has been developed. The ELISA method was checked with serum samples obtained from rabbits and pigs, and the parameters affecting the method were also analysed. ELISA plates were optimally coated with K88 antigen 0.5 microgram/ml for testing rabbit antiserum or with 1.25 microgram/ml for testing pig serum. Optimal concentrations of H202 (0.5%) and orthophenylene-diamine (OPD) (0.125%) were chosen when a 10-min incubation period was used. The expression of antibody levels as enzyme-immunosorbent units (EIU) significantly decreased the variability of results between duplicate plates, when compared with the expression of results as direct OD values. ELISA-K88 applied to a field study with serum samples from 141 vaccinated and 52 unvaccinated sows was shown to be useful in differentiating between samples from vaccinated and unvaccinated animals.

Fimbriae extracts from enterotoxigenic Escherichia coli strains of bovine and porcine origin with K99 and/or F41 antigens.

Fimbriae extracts obtained using the thermal shock method, from bovine and porcine enterotoxigenic Escherichia coli strains with K99 and/ or F41 antigens, were analyzed by SDS-PAGE, immunoblotting and haemagglutinating activity. Three major protein bands with molecular weights 17 kDa, 29.3 kDa and 30.9 kDa were detected depending on the strain assayed. A 17 kDa band was identified as the fimbrial subunit for K99 fimbriae and was detected in strains of bovine and porcine origin. The 30.9 kDa band was identified as the fimbriae subunit for F41 fimbriae and was detected in all porcine strains with F41 antigen and only in the bovine strains of serogroups O9 and 0101 that proved positive for F41 antigen. The 29.3 kDa band was shown to be antigenically related to F41 and K88, and was only detected in bovine strains of serogroups O8 (5 strains) and O20 (a single strain). We speculate that the 29.3 kDa band may be related to the CS31A antigen.

Serogroups of Escherichia coli isolated from piglets in Spain.

Serogroups of 1334 E. coli colonies isolated in Spain between 1986 and 1991 from piglets with diarrhoea, oedema disease and from healthy piglets, were determined. The serogroups determined in E. coli from diarrhoea were: O1, O2, O4, O5, O7, O8, O9, O12, O20, O21, O23, O25, O26, O32, O39, O45, O54, O75, O78, O80, O83, O91, O92, O101, O103, O113, O115, O116, O118, O125, O127, O138, O139, O141, O149, O153 and O157; from pigs with oedema disease: O8, O101, O138, O149 and O157; and from healthy piglets: O1, O2, O5, O7, O8, O9, O20, O21, O26, O29, O45, O64, O71, O80, O81, O91, O101, O105, O113, O115, O116, O126, O128, O132, O138, O139, O142, O146, O149, O152, O153, and O168. Serogroups O138, O141 and O149 were found to be statistically associated with enteric porcine colibacillosis (diarrhoea and oedema disease) (0.025 > P < 0.01). In addition, enterotoxigenic (ETEC) strains belonged to serogroups: O8, O9, O20, O101, O138, O141 and O149; verotoxigenic (VTEC) strains (VTEC) strains to serogroups 091 and 0138; and necrotizing (NTEC) strains to serogroups O2, O4, O8, O54, O78 and O83. Furthermore, 91.2% (249 out 273) of the ETEC (LT and/or STa) and/or VTEC strains belonged to only seven different serogroups. These major serogroups to which the ETEC and VTEC strains belonged, were determined in a lower percentage (21.2%) among non-toxigenic E. coli colonies isolated from sick and healthy piglets.

Toxigenic Escherichia coli in Spanish piggeries from 1986 to 1991.

Four-hundred and fourteen faecal samples from pigs with diarrhoea, oedema disease or healthy pigs, were collected from 65 piggeries located in different areas of Spain from 1986 to 1991. A total of 1334 Escherichia coli cultures were isolated from the pigs and studied for production of heat-labile (LT) and heat-stable (STa) enterotoxins, verotoxin (VT) and for type 1 (CNF1) and type 2 (CNF2) cytotoxic necrotizing factors. Strains producing enterotoxins (P < 0.001) or verotoxin (P < 0.05) were associated with enteric diseases of pigs. In the majority (82.3%) of piglets with strains, producing verotoxin the strains were also positive for production of STa enterotoxin. The most frequent toxin detected was STa. Although we isolated strains producing CNF1 from 1.5% of sick pigs, they were not statistically associated (P < 0.7) with enteric disease. Pigs may constitute a natural reservoir of CNF1 producing E. coli strains in Spain; their presence in the porcine intestine may be of significance in public health because such strains have been associated with human extraintestinal infections.

Serotypes and colonization factors of enterotoxigenic Escherichia coli isolated in various countries.

One hundred and six enterotoxigenic E. coli (ETEC) isolated from many geographical areas were serotyped and investigated for the presence of colonization factor antigens CFA/I and CFA/II, the expression of mannose-resistant haemagglutination (MRHA) and the levels of surface hydrophobicity. CFA/I was found in 6 (17%) of 36 LT+STa+ strains and in 15 (54%) of 28 STa+ strains; CFA/II was found in 16 (44%) of 36 LT+STa+ strains. None of 42 LT+ strains showed CFA/I or CFA/II. CFA/I was found in ETEC of serotypes O63:K-:H-, O78:K80, O128:K67 and O153:K:H45, whereas CFA/II was found in serotypes O6:H-, O6:K15:H16 and O6:K?:H40. Of the 69 CFA/I- CFA/II- ETEC strains, 9 (13%) showed MRHA with some of the seven erythrocyte species used and 21 (30%) were hydrophobic. Among the 21 hydrophobic strains CFA-negative we have detected: (i) 6 LT+ strains of serogroup O25 negative for MRHA, (ii) 5 strains O159 (4 LT+ and 1 LT+ STa+) also negative for MRHA, and (iii) 3 STa+ strains of serotype O27:K-:H7 that haemagglutinated calf and sheep erythrocytes when grown on Minca-Is. The 106 ETEC strains belonged to 20 different O serogroups. However, 77 (73%) were of one of nine serogroups (O6, O8, O25, O27, O78, O148, O153, O159 and O167). E. coli strains belonging to O6 and O153 groups predominated among ETEC isolated in Spain, O159 strains in the Central African Republic, O25 and O148 strains in Japan, and O15 and O78 strains in India.

Toxic and adhesive properties of Escherichia coli strains belonging to classic enteropathogenic serogroups.

Twenty-three strains belonging to classic enteropathogenic Escherichia coli (EPEC) serogroups were investigated for the production of heat-labile (LT) and heat-stable (STa) enterotoxins, verotoxins (VT), cytotoxic necrotizing factors CNF1 and CNF2, alpha-haemolysin (Hly), necrosis and modification of permeability in rabbit skin, lethal activity to mice, mannose-resistant (MRHA) and mannose-sensitive (MSHA) haemagglutination, relative cell surface hydrophobicity and the expression of P fimbriae. Of 23 EPEC strains, 7 (30%) belonging to serotypes O18ac: H7 (Hly+), O20: H26 (lethal), O26: H- (Hly+), O44: H18 (Hly+), O55: H- (CNF2+, necrotic and lethal), O119: H27 (VT+ and Hly+) and O142: H6 (lethal) produced toxic factors. Seven (30%) of 23 EPEC strains were MRHA+, 17 (74%) were MSHA+ and only 2 possessed high hydrophobicity. Two strains belonging to serotypes O18ac: H7 and O44: H18 that showed MRHA type IVa were fimbriated when grown on CFA medium.

Establishment of three categories of P-fimbriated Escherichia coli strains that show different toxic phenotypes and belong to particular O serogroups.

Eight hundred and nineteen strains of Escherichia coli isolated in Spain between 1986 and 1991 from extraintestinal infections and feces of healthy controls were investigated for expression of P-fimbriae using a particle agglutination test. Among strains causing urinary tract infections, sepsis and other extraintestinal infections, P-fimbriae were found in 31% (130/420) (P < 0.001), 25% (30/118) (P < 0.001) and 12% (11/92) (P < 0.5) respectively. In contrast, only 7% (14/189) of faecal isolates from healthy individuals carried P-fimbriae. According to two more common toxic markers detected in this study (alpha-haemolysin and cytotoxic necrotizing factor type 1), P-fimbriated E. coli strains were grouped into three categories: haemolysin+cytotoxic necrosing factor+ (Hly+CNF1+) (68/185; 37%), haemolysin+cytotoxic necrosing factor- (Hly+CNF1-) (61/185; 33%) and Hly-CNF1- (56/185; 30%). The 185 P-fimbriated strains belonged to 17 different O serogroups. However, 148 (80%) were of one of six serogroups (O1, O2, O4, O6, O7 and O18). The most frequent serogroups determined in the Hly+CNF1+ strains were the O4 and O6 (53/68; 78%), in the Hly+CNF1- strains it was the O18 (27/61; 44%) and in the Hly-CNF1- strains the O1, O2 and O7 (41/56; 73%). The majority (160/185; 86%) of P-fimbriated E. coli expressed the mannose-resistant haemagglutinin type IVa.

Characteristics of haemolytic Escherichia coli with particular reference to production of cytotoxic necrotizing factor type 1 (CNF1).

A total of 1,106 Escherichia coli strains isolated in Spain between 1986 and 1991 from extraintestinal infections and faeces of healthy controls were examined for production of alpha-haemolysin (Hly). Among strains causing urinary tract infections, sepsis and other extraintestinal infections, Hly production was detected in 51% (P < 0.001), 32% (P < 0.001) and 18% (P < 0.02), respectively. In contrast, only 9% of faecal isolates from healthy individuals synthesized Hly. The 356 haemolytic E. coli strains characterized in this study belonged to 28 different serogroups. However, 284 (80%) were of one of eight serogroups (02, 04, 06, 08, 018, 022, 075 and 083); 40% and 31% of haemolytic strains expressed P fimbriae and mannose-resistant haemagglutination (MRHA) type III, respectively. We have found that haemolytic isolates of E. coli may clearly be divided into two categories on the basis of the ability to produce cytotoxic necrotizing factor type 1 (CNF1). The serogroups and adhesins determined in Hly+CNF1+ strains were generally different from those found in Hly+CNF1- strains. Thus, serogroups 02, 06 and 075 were associated with haemolytic E. coli producing CNF1+, whereas serogroups 01, 08, 018, 028 and 086 were established more frequently among Hly+CNF1- strains. While expression of P fimbriae was more frequently detected in Hly+CNF1- strains (70 versus 29%, P < 0.001), MRHA type III was usually identified in Hly+CNF1+ E. coli (42 versus 1%, P < 0.001). Furthermore, the sonic extracts of Hly+CNF1+ strains caused necrosis in rabbit skin (96 versus 25%, P < 0.001) and death in intraperitoneally injected mice (73 versus 11%, P < 0.001) more frequently than sonic extracts of Hly+CNF1- strains.

Serogroups of Escherichia coli strains producing cytotoxic necrotizing factors CNF1 and CNF2.

The serogroups of 396 necrotizing Escherichia coli of human and bovine origin isolated in Spain between 1979 and 1991 have been determined. The 270 cytotoxic necrotizing factor strains belonged to 22 different O serogroups; however, 84% (226 of 270) were of one of seven serogroups (O2, O4, O6, O14, O22, O75 and O83). Although necrotizing E. coli producing cytotoxic necrotizing factor 2 belonged to 28 different serogroups, only six of them (O1, O3, O15, O55, O88 and O123) accounted for 60% (76 of 126) of cytotoxic necrotizing factor 2 strains. Furthermore, only 3% (4 of 126) of cytotoxic necrotizing factor 2 strains belonged to serogroups most common among strains producing cytotoxic necrotizing factor 1. The majority of necrotizing E. coli producing cytotoxic necrotizing factor 1 were obtained from human extraintestinal infections, whereas cytotoxic necrotizing factor 2 strains were isolated from stools of healthy and diarrhoeic calves.

[Escherichia coli virulence factors causing peritonitis, appendicitis and other extraintestinal infections]. / Factores de virulencia de Escherichia coli causantes de peritonitis, apendicitis y otras infecciones extraintestinales.

Fifty-nine E. coli strains isolated from clinical cases of peritonitis, appendicitis, cholecystitis, wounds and respiratory infections as well as from other miscellaneous sources were investigated. A control group constituted by 475 E. coli strains isolated from the faeces of healthy individuals were also studied. E. coli O-grouped and investigated for production of cytotoxic necrotizing factor CNF1 and alpha-haemolysin (Hly), expression of P fimbriae and mannose-resistant (MRHA) and mannose-sensitive (MSHA) haemagglutination. Virulence factors significantly associated with extraintestinal strains were: production of CNF1 (19% versus 5%, p < 0.001), Hly (27% versus 9%; p < 0.001) and expression of MRHA (44% versus 16%; p < 0.001). The majority of extraintestinal strains (68% versus 36%; p < 0.001), in contrast with faecal E. coli, belonged to O serogroups frequently detected in uropathogenic and bacteraemic E. coli. These results suggest that E. coli causing different types of extraintestinal infections show similar virulence factors and belong to the same serogroups. However, between E. coli isolated from intraabdominal, wound and respiratory infections the number of strains with virulence factors was lower than in E. coli causing urinary tract infections and sepsis.

Enterotoxigenic and necrotizing Escherichia coli in human diarrhoea in Spain.

Enterotoxigenic Escherichia coli (ETEC) strains of serotype 0153: K-:H45 CFA/I+ STa+ were associated with two outbreaks of neonatal diarrhoea that occurred in two different hospitals of Madrid, in one of which several children died. Two other outbreaks were associated with ETEC strains of serotypes 0159: K-:H21 (LT+) and 0159: K-:H4 (LT+ STa+) without CFA/I and CFA/II colonization factors. Necrotizing E. coli (NTEC) strains of serotype 06:K13, producing the cytotoxic necrotizing factor CNF1 and alpha-haemolysin, were also associated with two outbreaks of neonatal diarrhoea that occurred in a hospital in Madrid and in a hospital in Talavera de la Reina. The results of the characterization of some ETEC and NTEC strains isolated from sporadic cases of diarrhoea are also discussed.

Bovine Escherichia coli of serotypes O55:H4 and O55:H21 which produce CNF2 express P fimbriae and Vir surface antigen, respectively.

We have characterized the toxic and adhesive properties of Escherichia coli strains producing the second type of cytotoxic necrotizing factor (CNF2) and belonging to the classic enteropathogenic serogroup O55. Bovine CNF2 strains of serotype O55:H4 express P fimbriae as do pyelonephritic Escherichia coli that cause infections in humans. In contrast, strains of serotype O55:H21 which produce CNF2 from bovine origin possess the Vir surface antigen. One human strain of serotype O55:H- was positive for production of CNF2, but was negative for the two adhesive factors and for mannose-resistant haemagglutination.

Enterotoxins, colonization factors and serotypes of enterotoxigenic Escherichia coli from humans and animals.

Enterotoxigenic Escherichia coli (ETEC) strains may synthesize both thermolabile (LT-I and LT-II) and thermostable (STa and STb) enterotoxins. Whereas thermolabile enterotoxins are high molecular weight proteins (85,000 d-90,000 d) composed by a single enzymatic A subunit combined with five B subunits which enable toxin for the receptor recognition, thermostable enterotoxins are small peptide chains with molecular weight between 1,900 d and 5,000 d. In addition to the synthesis of enterotoxins, the ability of ETEC strains to cause diarrhoea is also conditioned by the possession of colonization factors which enable bacteria adhere-to and colonize the luminal surface of small bowel. Colonization factors in ETEC strains were located in rigid fimbriae and flexible fibrils constituted by protein subunits ranging in size from 14,500 d to 31,000 d and usually responsible for mannose-resistant haemagglutination with determined erythrocyte species. Both enterotoxins and colonization factors are controlled by plasmids. There exist plasmids which may code separately enterotoxins and colonization factors, and besides there also exist recombinant plasmids coding together these two virulence factors. Human ETEC strains may synthesize LT-I and/or STa enterotoxins, they may possess the colonization factors named CFA/I, CFA/II, CFA/III or CFA/IV, and they belong mainly to serogroups O6, O8, O15, O20, O25, O27, O63, O77, O78, O114, O115, O126, O128, O139, O148, O153, O159 and O167. ETEC strains from porcine origin synthesize LT-I, STa and/or STb, they possess the colonization factors K88, P987, K99 or F41, and they usually belong to serogroups O8, O9, O20, O45, O64, O101, O115, O138, O141, O147, O149 and O157. Bovine and ovine ETEC strains are usually STa producers harbouring on the bacterial surface K99 or F41 colonization factors and they belong to serogroups O8, O9 and O101. Nevertheless, some particular bovine ETEC strains synthesizing LT-II have been described. Thus, a high specificity level between ETEC strains causing diarrhoea in humans and domestic animals can be observed. This is mainly due to the specific recognition between bacterial colonization factors and the epithelial receptors during host-parasite interaction.

Enterotoxigenic Escherichia coli associated with infant diarrhoea in Galicia, north-western Spain.

To assess the role of enterotoxigenic Escherichia coli (ETEC) in infantile diarrhoea, 482 children with diarrhoea and 103 healthy controls, from three localities of Galicia, north-western Spain, were investigated between 1985 and 1988. Rotavirus (37.3%) and Salmonella spp. (12.8%) were the most common causal agents, followed by ETEC (3.9%), Campylobacter jejuni (2.3%), Shigella spp. (0.9%) and Yersinia enterocolitica (0.5%). ETEC were significantly more frequently isolated from children with diarrhoea who were under 1 month of age (26.5%) than from older diarrhoeic children (2.2%) (p less than 0.001) or from healthy children who were under 1 month of age (0%) (p less than 0.05). Among children who harboured ETEC, five of the nine children under 1 month of age developed diarrhoea in hospital, whereas none of the 10 children over 1 month of age did so. Seventeen ETEC isolates produced heat-stable enterotoxin (STa) only, four produced only heat-labile enterotoxin (LT), and two produced both toxins. Colonisation factor antigens CFA/I and CFA/II were detected in 11 (55.0%) of the 20 ETEC isolates that remained enterotoxigenic after maintenance in the laboratory. Most ETEC isolates belonged to serotypes O153:K-:H45 (nine STa+ CFA/I+ isolates), O27:K-:H7 (three STa+ isolates) or O6:K15:H16 (two LT+ STa+ CFA/II+ isolates). Our results suggest that ETEC constitute an important cause of neonatal diarrhoea in this part of Spain.