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1.
DNA gene probes specific for genes encoding heat labile enterotoxin (LTI), heat stable enterotoxins (STIa, STII), vero cytotoxins (VT1, VT2), and adhesins K88 (F4), K99(F5), F41 and 987P(F6) were used to examine 873 isolates of E. coli from cases of diarrhoea (680 from pigs, 187 from cattle and six from sheep). A total of 188 were toxin gene positive and of these 84 belonged to the classical ETEC serogroups. Of the other 104 toxin gene positive strains, 80 hybridized with the VT2 probe of which 34 were from cases of porcine post-weaning diarrhoea belonging to serogroup 0138:K81 and 22 were untypable strains from cattle.  相似文献   

2.
Oligonucleotide primers were designed for the specific polymerase chain reaction (PCR) amplification of the enterotoxins STIa and LTI and of the verocytotoxins VT1 and VT2. All of 184 E. coli isolates from cases of diarrhoea from pigs, cattle and sheep gave identical toxin profiles by PCR and gene probe. Differentiation between VT2 and VT2v was achieved using two oligonucleotide primers pairs in PCR and showed that all of 34 VT2+ porcine isolates, of which 23 were 0138:K1, harboured VT2v whereas 20 VT2+ bovine and ovine isolates harboured VT2. No isolate harboured both VT2 polymorphs. Simplified methods for sample preparation for PCR were examined and showed that PCR was not inhibited by direct addition of broth culture to the reaction mixture.  相似文献   

3.
The presence of fimbrial adhesin F18 is frequently found in enterotoxigenic Escherichia coli (ETEC) and verotoxigenic E. coli (VTEC) strains responsible for diarrhoea and oedema disease of weaned pigs. The F18 adhesin occurs in two antigenic variants: F18ab is characteristic of VTEC while F18ac is more typical for ETEC. F18 encoding plasmids of 17 phenotypically characterized porcine E. coli isolates (10 ETEC, 6 VTEC and 1 ETEC/VTEC) were tested with a DNA probe for F18 fimbrial adhesin and with replicon probes for the RepFIa, RepFIb and for the RepFIc family of basic replicons. In all the cases, the F18 probe hybridized to only one plasmid band of size higher than 42MDa. All F18 plasmids were determined to be unireplicon plasmids belonging to the RepFIc replicon family of the F incompatibility complex. There was no difference between F18ac plasmids of ETEC and F18ab plasmids of VTEC strains in terms of replicon type or subtype. However, the size of F18ab plasmids of the VTEC strains varied between 42 and 98MDa, in contrast to F18ac plasmids of ETEC strains (constantly approximately 98MDa).  相似文献   

4.
Samples of faeces from 57 dogs with acute diarrhoea, 82 dogs with chronic diarrhoea, 34 clinically healthy household dogs and 88 kennelled control dogs were analysed by hybridisation, using DNA probes to detect enteropathogenic Escherichia coli (EPEC) and enterotoxigenic E coli (ETEC), verocytotoxin-producing E coli (VTEC), enterohaemorrhagic E coli (EHEC), enteroinvasive E coli (EIEC) and enteroaggregative E coli (EAggEC). Samples of duodenal juice from 60 of the 82 dogs with chronic diarrhoea were also examined. Significantly more of the dogs with diarrhoea were excreting EPEC (acute 35.1 per cent, chronic 31.7 per cent) and VTEC (acute 24.6 per cent, chronic 28 per cent) than the kennelled dogs (EPEC 17.1 per cent, VTEC 0 per cent) or the household control dogs (EPEC 6 per cent, VTEC 5.9 per cent). Enteropathic E coli was also detected in the duodenal juice of 23 of 60 (38.3 per cent) of the dogs with chronic diarrhoea. The EPEC attaching and effacing A (eaeA) gene and the verocytotoxin 1 (VR1) gene coding for VTEC were often found together. There was good agreement between in vitro studies and hybridisation for the detection of eaeA and VT1. Isolates from the dogs with diarrhoea adhered significantly more to Hep-2 cells, and VT1-positive strains from the dogs with diarrhoea consistently killed more than 50 per cent of Vero cells.  相似文献   

5.
Shiga/verocytotoxins and Shiga/verotoxigenic Escherichia coli in animals.   总被引:6,自引:0,他引:6  
J Mainil 《Veterinary research》1999,30(2-3):235-257
Vero/Shiga toxins (VT/Stx) have an A-B structure: the A subunit carries the enzymatic activity and the B subunit binds the toxin to the membrane receptor (Gb3 or Gb4). The VT/Stx inhibit protein synthesis in the target eucaryotic cells, mainly the endothelial cells of blood vessels. The VT/Stx are subdivided into two families. VT1/Stx1 is a homogeneous family of toxins identical to the Stx of Shigella dysenteriae. VT2/Stx2 is a more heterogeneous family of toxins more distantly related to this Stx toxin. The VT2/Stx2 variants can be distinguished by the polymerase chain reaction (PCR) and/or the reaction with monoclonal antibodies. The VT/Stx-producing Escherichia coli are also subdivided into two main groups on the basis of the presence or absence of additional properties: the enterohaemorrhagic E. coli (EHEC) induce the formation of attaching/effacing lesions and carry a 60 MD plasmid encoding a specific haemolysin (the enterohaemolysin); the vero/shiga-toxigenic E. coli (VTEC/STEC) do not show these properties. The EHEC are isolated from humans and ruminants, especially young calves. They are associated with haemorrhagic enterocolitis and its sequelae in humans, the haemolytic-uraemic syndrome (HUS). The VT/Stx play a role in the occurrence of blood in the faeces and in the HUS by their action on the endothelial cells of blood vessels in the intestinal submucosa and in the renal glomeruli, after resorption through the intestinal walls. The VTEC/STEC are isolated from piglets, calves and humans. In recently weaned piglets, they cause the oedema disease, an enterotoxaemia characterized by subcutaneous, mesenteric and cerebral oedemas, with nervous disorders as main clinical signs. The oedema disease is the consequence of the action of the VT/Stx on the endothelial cells of blood vessels in various organs. In calves and humans, the role in disease of VTEC/STEC is controversial, but they could be associated with some cases of diarrhoea and HUS. The case of the O157:H7 EHEC which are present in healthy cattle of various ages, but are highly virulent for humans is of special interest. The potential zoonotic aspect of VT/Stx-producing E. coli infections in animals is detailed chapter by chapter. Prophylaxis of these infections by vaccination is the subject of the discussion on the future of the research studies on these pathogenic bacteria.  相似文献   

6.
Cheng D  Sun H  Xu J  Gao S 《Veterinary microbiology》2005,110(1-2):35-39
F18ab and F18ac are important fimbrial colonization factors of verotoxigenic Escherichia coli (VTEC) and/or enterotoxigenic E. coli (ETEC) in weaned piglets with edema disease and/or diarrhea. To further investigate their prevalence and correlation to pathogenic E. coli, a duplex PCR, using three primers derived from the nucleotide sequence of the F18 major fimbrial subunit gene (fedA), and a direct agglutination test, using a monoclonal antibody specific for the antigenic factor 'a' of F18, were performed. Among 60VTEC, 24VTEC/ETEC and 24 ETEC isolates tested from weaned piglets with edema disease and/or diarrhea in different pig farms in the Jiangsu Province of China, 52 isolates (48.15%) were positive in the direct agglutination test and 63 isolates (58.33%) were positive in the duplex PCR. Among 63 PCR-positive isolates, 53 isolates (49.07%) were F18ab-positive and 10 isolates (9.26%) were F18ac-positive. In addition, the F18ab gene was more frequently detected in VTEC (61.67%) or VTEC/ETEC (62.50%) than in ETEC (4.17% only), while the F18ac gene was more frequently detected in VTEC/ETEC (33.33%) than in ETEC (8.33%) or VTEC (0%). Furthermore, F18ab was more frequently associated with Shiga toxin 2e (Stx2e), whereas F18ac was more frequently associated with enterotoxin ST I. These results suggest that the duplex PCR performed in this experiment is a more reliable method for identification of F18+E. coli, and that F18 is a more important virulence factor of VTEC and VTEC/ETEC.  相似文献   

7.
The study comprises fifty 4 to 12 weeks old pigs that died from oedema disease or severe diarrhoea. Smears were prepared from the mucosa of duodenum, jejunum and ileum, and by immunofluorescence F107 fimbrial antigens were detected. E. coli. strains were isolated from the intestines and were characterised by slide agglutination (serogroup and F107 fimbriae production), by their cytotoxicity for Vero cells, and by gene amplification (genes coding for the major F107 subunit FedA, the toxin causing oedema disease SLT-IIv, and enterotoxins LTI, STIa and STII). F107 fimbriae were demonstrated in association with E. coli of serogroups O139:K12 and O141:K85a,b but not of serogroup O149:K91:F4a,c. Expression in culture of F107 fimbriae by some isolates gave additional evidence for production of these fimbriae by ETEC strains. The genetic determinant of SLT-IIv was found in association with F107, and could not be detected in serogroup O149:K91:F4a,c. Gene fedA was demonstrated in two isolates which were devoid of SLT-IIv. Most isolates from cases of oedema disease belonged to serogroup O139:K12 and did not contain enterotoxin genes. Isolates from pigs that suffered from diarrhoea were serotyped O141:K85a,b or O149:K91:F4a,c, and carried at least two enterotoxin genes in their genomes. In a small proportion of the cases F107 antigens were demonstrated in intestinal smears although gene fedA was not detected in the corresponding isolates. The results confirm the importance of F107 fimbriae as virulence factor in oedema disease E coli strains, but also demonstrate that F107 fimbriae can be found in association with postweaning diarrhoea isolates. In these latter strauns enterotoxins were always demonstrated, irrespective of the presence of toxin SLT-IIv.  相似文献   

8.
A novel common colonization factor was detected in enterotoxigenic strains of Escherichia coli isolated from intestinal contents of piglets affected with postweaning diarrhoea. This factor was antigenically distinct from the previously described K88, K99, F41, 987P, CFAI, CFAII and Att25 fimbrial antigens. E. coli strains possessing this factor adhered to the pig intestinal brush borders and one strain, used in experimental infection in weanlings, colonized the intestinal epithelium and induced diarrhoea. Examination of 212 toxigenic strains of E. coli isolated from weanlings revealed the presence of the novel common colonization factor in 83 strains, belonging to serogroups O25, O108, O138, O141, O147 and O157. The antigen K88 was detected in 47 strains belonging to serogroups O8, O141, O147 and O149.  相似文献   

9.
A cross-sectional study was conducted to determine the prevalence and characteristics of verocytotoxigenic Escherichia coli (VTEC) on 25 dairy farms each located in Waller field and Carlsen field farming areas in Trinidad. On each selected farm, faecal samples were collected from milking cows, calves and humans; rectal swabs were obtained from pet farm dogs; bulk milk was sampled as well as effluent from the milking parlour. Escherichia coli was isolated from all sources on selective media using standard methods. Isolates of E. coli were subjected to slide agglutination test using E. coli O157 antiserum, vero cell cytotoxicity assay to detect verocytotoxin (VT) and heat labile toxin (LT) production, the polymerase chain reaction (PCR) to detect VT genes, and the dry spot test to screen for E. coli O157 and non-O157 strains. In addition, faecal samples from animal and human sources were tested for VT genes using PCR. Of a total of 933 E. coli isolates tested by the slide test, eight (0.9%) were positive for the O157 strain. The vero cell cytotoxicity assay detected VT-producing strains of E. coli in 16.6%, 14.6%, 3.2% and 7.1% of isolates from cows, calves, farm dogs and humans respectively (P < 0.05; chi(2)). For LT production, the highest frequency was detected amongst isolates of E. coli from calves (10.8%) and the lowest (0.0%) amongst isolates from humans and bulk milk (P < 0.05; chi(2)). Of the 61 VT-producing isolates by vero cell cytotoxicity assay tested by PCR, the VT, LT and eae genes were detected in 62.3%, 4.9% and 1.6% respectively (P < 0.05; chi(2)). Amongst the 45 E. coli isolates that were VT positive (vero cell) or VT-gene positive by PCR, 2.2%, 2.2%, 4.4% and 6.7% belonged to non-O157 strains O91, O111, O103 and O157, respectively, as determined by the Dry spot test. Detection of VTEC strains in milk and dairy animals poses a health risk to consumers of milk originating from these farms. In addition, the demonstration of VTEC strains in humans, VT gene in faecal samples and E. coli isolates as well as non-O157 VTEC strains of E. coli are being documented for the first time in the country.  相似文献   

10.
Cattle arriving for slaughter at a large abattoir in northern Italy between April 1997 and January 1998 were examined for intestinal carriage of Verocytotoxin-producing Escherichia coli (VTEC) O157 using an immunomagnetic separation technique. Sixty sorbitol non-fermenting VTEC O157 strains were isolated from 59 (13.1%) of the 450 cattle examined. In particular, VTEC O157 was found in 37 (16.6%) of 223 feedlot cattle and in 22 (16.1%) of 137 dairy cull cows, but not in the 90 veal calves sampled. The isolation rate was higher during warm weather (17.5%), falling to an average of 2.9% during the winter months. VT-negative, O157 latex-agglutinating E. coli strains were isolated from 23 (5.1%) of the 450 animals. PCR analysis showed that all 60 VTEC O157 strains carried the VT2 gene and that 25 strains also carried the VT1 gene. In addition, four of the VT-negative, O157 latex-agglutinating E. coli strains carried the VT2 gene. Atypical biochemical features were observed in some VTEC O157: two strains (3.3%) showed beta-glucuronidase activity, and seven (11.7%) produced urease.  相似文献   

11.
We tested hemolytic E. coli from 86 pigs with edema disease or colidiarrhea. They were tested serologically and with nonradioactive digoxigenin-dUTP labelled probes for the presence of enterotoxin or Shiga-like-toxin genes. By slide-agglutination we detected 38 cases with E. coli O149:K88, 28 with E. coli O139:82B and 20 with E. coli O141. E. coli of serogroup O149:K88 isolated from diarrheic pigs, reacted with the probes for LT and STb genes. Edema disease E. coli O139:82B reacted with the SLTII probe. E. coli O141, isolated from colidiarrhea or edema disease showed a diversity of toxin gene patterns. All the E. coli O141 from diarrheic pigs reacted with the probes for LT and STap in addition to SLTII. No strains isolated from pigs with edema disease possessed any of these enterotoxin genes. Gene probe technique confirmed the serological method as useful tool for diagnosing E. coli O149:K88 and O139:82B as ETEC or VTEC, respectively. On the other hand only the demonstration of toxin genes with probes could explain the pathological findings in the pigs shedding E. coli of serogroup O141.  相似文献   

12.
利用F18菌毛a因子单克降抗体以及已建立的鉴定F18菌毛及其亚型的双重PCR法,对来自断奶仔猪水肿病和/或腹泻病例的60株VTEC、24株VTEC/ETEC以及24株ETEC的进行了F18菌毛检测,以了解F18ab^+和F18ac^+大肠杆菌在江苏省断奶仔猪群的分子流行病学。结果表明:通过F18菌毛a因子单克隆抗体,可检测出52株大肠杆菌为F18^+,检出率为48.15%;而通过双重PCIL方法,共检测出63株大肠杆菌为F18^+,检出率为58.33%,其中53株(49.07%)为F18ab^+10株(92.6%)为F18ac^+。另外还发现:在VTEC、VTEC/ETEC以及ETEC的菌株之间,这2种F18菌毛亚型的分子流行病学是不同的。在VTEC中,F18ab^+,菌株37株(61.67%),未发现F18ac^+菌株;在VTEC/ETEC中,F18ab^+菌株15株(62.50%),F18ac^+菌株8株(33.33%);而在ETEC中F18ab^+菌株只有1株(4.17%),F18ac^+菌株只有2株(8.33%)。以上数据表明:④PCR法检测F18菌毛优于单抗法;②F18菌毛是VTEC/ETEC、VTEC的重要致病因子,而在ETEC中则明显低于VTEC/ETEC和VTEC;⑧F18ab^+菌株一般为SLT-IIe^+,而F8ac^+菌株一般为STI^+。  相似文献   

13.
World-wide, enterotoxigenic Escherichia coli (ETEC) and verotoxigenic E. coli (VTEC)-induced diarrhea are economically important for porcine producers. Our aim was to investigate the prevalence of toxin and fimbrial genes among E. coli isolated from diarrheic piglets from randomly selected piggeries in Zimbabwe.We used multiplex PCR for screening STa, STb, LT, and Stx-2e toxins. Subsequently F4, F5, F6, F18 and F41 fimbriae genes were screened in toxin positive isolates. Toxin positive strains lacking tested fimbriae genes were characterized using transmission electron microscopy, agglutination and agglutination inhibition tests. Approximately 32% of the 1,984 isolates tested positive for STa, STb, LT or Stx-2e genes. Of these, approximately 81% had F4, F5, F6, F18 or F41 fimbriae genes. The remaining toxin positive strains lacked tested fimbriae genes and appeared to either express F1-like fimbriae, or lacked fimbriae. The data constitute an important framework for implementation of prevention measures, such as using relevant fimbriae-based vaccines against ETEC induced diarrhea or VTEC-induced edema.  相似文献   

14.
Three strains of enterotoxigenic Escherichia coli (ETEC) (064:KSNT, K88ac; 020:KSNT, K88ac and 08:K85ab, K99) originally cultured from outbreaks of diarrhoea in piglets a few hours old, were administered orally to gnotobiotic piglets. There was a marked age-related difference in the clinical response to infection between the 3 strains although they all produced heat-stable toxin. All 3 strains produced severe clinical signs of depression, anorexia, vomiting, diarrhoea, followed by dehydration and death in one-day-old piglets. In piglets infected at 3 days of age the two K88+ ETEC caused diarrhoea and death but the K99+ ETEC induced moderate diarrhoea only. In piglets infected at 7 days of age, the 064 strain produced severe diarrhoea and death, and 020 strain caused mild diarrhoea in 3 of 6 piglets with one death while the 08 strain caused no illness. Pathological changes in the intestinal tract associated with these infections were minimal, or absent. Immunofluorescent staining with homologous hyperimmune sera demonstrated adherence of the 3 ETEC strains to the brush border of small intestinal epithelial cells. Fluorescing organisms were observed in all infected piglets irrespective of the severity of clinical signs but the degree and extent of colonisation varied with the age of the piglets and the infecting strain. This may explain the difference in clinical response between the 3 strains.  相似文献   

15.
The presence of the astA gene responsible for production of enteroaggregative Escherichia coli heat-stable enterotoxin 1 (EAST1) was examined in E. coli strains isolated from pigs with postweaning diarrhoea. Two hundred and seven isolates were tested using PCR for the astA marker and for heat-labile I (LTI), heat-stable I (STI), and heat-stable II (STII) enterotoxin genes. Moreover, the isolates were also analysed for their serotypes (O and K antigens) as well as for fimbrial adhesins using agglutination methods. It was shown that 96 (46.4%) of the isolates possessed the astA genetic determinant. The most common EAST1-positive E. coli serotype was O149:K91 and these strains were mostly LTI/STII-positive. A close correlation between the presence of F4 fimbriae and the EAST1 gene was also observed: 88 of 96 (91.7%) astA(+) isolates tested possessed the F4 antigen. Thus, EAST1 enterotoxin may represent an additional virulence determinant playing a role in the pathogenesis of porcine colibacillosis.  相似文献   

16.
The outcome of experimental intestinal infections with enterotoxigenic Escherichia coli (ETEC) is dependent on several factors. An important factor is adhesion of the challenge strain to the intestinal mucosa. The test for susceptibility towards ETEC adhesion has so far been made by an intestinal adhesion test made after slaughter of piglets. However, in an experimental infection study with the purpose to obtain diarrhoeic piglets, it would be an advantage to test for susceptibility prior to experimentation. The Mucin 4 gene on porcine chromosome 13 has been proposed as a candidate gene for the production of the specific ETEC F4ab/ac receptor, and a DNA marker-based test has been developed to allow genotyping for ETEC F4ab/ac resistance/susceptibility [J?rgensen, C.B., Cirera, S., Archibald, A.L., Anderson, L., Fredholm, M., Edfors-Lilja, I., 2004. Porcine polymorphisms and methods for detecting them. International application published under the patent cooperation treaty (PCT). PCT/DK2003/000807 or WO2004/048606-A2]. The aim of this study was to test an experimental model for ETEC O149:F4ac-induced diarrhoea in piglets, selected for susceptibility towards ETEC O149:F4ac adhesion prior to experimentation using a DNA marker-based test. Sixty-two healthy 25-32 days old recently weaned Danish crossbred piglets were used. All piglets were tested prior to experimentation for susceptibility or resistance towards ETEC O149:F4ac adhesion. Thirty-nine piglets, both susceptible and resistant, were oro-gastric intubated with 10(9)CFU of ETEC O149:F4ac and 23 age-matched piglets, both susceptible and resistant, were used as non-infected controls. Of susceptible piglets, challenged with ETEC O149:F4ac, 74% had ETEC O149:F4ac-associated diarrhoea first day after first challenge, which were significantly higher relatively to the resistant and challenged piglets where 20% had diarrhoea (p=0.04). This study suggests a model for experimental ETEC induced diarrhoea.  相似文献   

17.
The adsorption property of activated charcoal on verotoxin (VT)-producing Escherichia coli (VTEC) was examined using E. coli O157:H7. In the present study, E. coli O157:H7 strains were effectively adsorbed by activated charcoal. Adsorption was dose-dependent, and the maximum adsorption occurred within 5 min. At 10 mg of activated charcoal, bacteria tested were completely adsorbed. Activated charcoal also had the capacity to adsorb toxin (verotoxin 2) activity from the bacterial extract. Furthermore, the adsorption efficiency of activated charcoal for the normal bacterial flora in the intestine was assessed using Enterococcus faecium, Bifidobacterium thermophilum, and Lactobacillus acidophilus. Activated charcoal showed lower binding capacity to the normal bacterial flora tested than that to E. coli O157:H7 strains. These results suggest that activated charcoal could be a good adsorbent system for the removal of VTEC and verotoxin.  相似文献   

18.
A multiplex polymerase chain reaction (PCR) system was developed for identification of enterotoxigenic Escherichia coli (ETEC) strains and to differentiate them from other gram negative enteric bacteria. This test simultaneously amplifies heat-labile (LTI) and heat-stable (STI and STII) toxin sequences and the E. coli-specific universal stress protein (uspA). The specificity of the method was validated by single PCR tests performed with the reference E. coli and non-E. coli strains and with bacteria isolated from pig feces. The multiplex PCR allowed the rapid and specific identification of enterotoxin-positive E. coli and may be used as a method for direct determination of ETEC and to differentiate them from other E. coli and gram-negative enteric isolates.  相似文献   

19.
The objectives of this study were to determine the prevalence of enteric verocytotoxigenic E. coli (VTEC) infection in a population of cats in Ontario, and to determine whether an association exists between the presence of VTEC and feline diarrhea. Fecal samples from 179 cats, representing 113 cats with diarrhea and 66 cats with normal feces, were cultured for E. coli. The fecal cultures were screened for verocytotoxin activity with a Vero cell assay. Confirmation of the presence of verocytotoxin (VT) genes was done with polymerase chain reaction (PCR) amplification; the frequency of occurrence of the genes for generic VT, VT1, and VT2 was determined. VTEC-positive samples were defined as those that demonstrated cytotoxicity on the Vero cell assay and yielded E. coli possessing one or more of the VT genes. All VTEC-positive isolates were serotyped. The overall prevalence of enteric VTEC infection in the cats was 12.3% (22/179). Statistical analysis of the case-control data showed no significant association between VTEC infection and diarrheal illness. The majority of the cats with VT-positive E. coli were positive for the presence of the generic VT, rather than for VT1 or VT2; it is therefore possible that a novel verocytotoxin gene may exist in E. coli isolated from cats. Eight VTEC strains were identified by serotyping; 4 of these serotypes have previously been isolated from humans, and 2 from cattle, suggesting that cats may be capable of acting as reservoirs for human and bovine VTEC serotypes.  相似文献   

20.
猪水肿病大肠杆菌分离、鉴定及药敏试验   总被引:6,自引:3,他引:3  
本实验从疑似猪水肿病的病例分离到5株大肠杆菌,O抗原鉴定结果表明所有菌株均为O139血清型;应用F18ab菌毛单克隆抗体对这5株大肠杆菌能否表达F18ab菌毛进行了鉴定,结果表明其中2个菌株能表达F18ab菌毛;利用聚合酶链式反应(PCR)对志贺氏菌样毒素Ⅱ型变异体(SLT-Ⅱe)操纵子基因保守区进行了扩增,结果发现在能表达F18ab菌毛2个菌株中可扩增一段特异性序列。以上数据表明这2株大肠杆菌为致仔猪水肿病大肠杆菌。药敏试验表明这两株菌株均对氟哌酸、妥布霉素、庆大霉素、利福平等抗生素高度敏感。  相似文献   

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