Development of Similar Broth Microdilution Methods to Determine the Antimicrobial Susceptibility of Flavobacterium columnare and F. psychrophilum

Flavobacterium columnare and F. psychrophilum are major fish pathogens that cause diseases that may require antimicrobial therapy. Choice of appropriate treatment is dependent upon determining the antimicrobial susceptibility of isolates. Therefore we optimized methods for broth microdilution testing of F. columnare and F. psychrophilum to facilitate standardizing an antimicrobial susceptibility test. We developed adaptations to make reproducible broth inoculums and confirmed the proper incubation time and media composition. We tested the stability of potential quality-control bacteria and compared test results between different operators. Log phase occurred at 48 h for F. columnare and 72–96 h for F. psychrophilum, confirming the test should be incubated at 28°C for approximately 48 h and at 18°C for approximately 96 h, respectively. The most consistent susceptibility results were achieved with plain, 4-g/L, dilute Mueller–Hinton broth supplemented with dilute calcium and magnesium. Supplementing the broth with horse serum did not improve growth. The quality-control strains, Escherichia coli ATCC 25922 and Aeromonas salmonicida subsp. salmonicida ATCC 33658, yielded stable minimal inhibitory concentrations (MIC) against all seven antimicrobials tested after 30 passes at 28°C and 15 passes at 18°C. In comparison tests, most MICs of the isolates agreed 100% within one drug dilution for ampicillin, florfenicol, and oxytetracycline. The agreement was lower with the ormetoprim–sulfdimethoxine combination, but there was at least 75% agreement for all but one isolate. These experiments have provided methods to help standardize antimicrobial susceptibility testing of these nutritionally fastidious aquatic bacteria.

Received June 24, 2015; accepted October 2, 2015.     

Improved Broth Microdilution Method for Antimicrobial Susceptibility Testing of Francisella Noatunensis Orientalis

In this project we optimized a minimal inhibitory concentration testing protocol for Francisella noatunensis orientalis. Thirty-three F. noatunensis orientalis isolates recovered from different fish species and locations were tested, and Escherichia coli ATCC 25922 was used as a quality control reference strain. A modified cation-adjusted Mueller Hinton broth supplemented with 2% IsoVitalex and 0.1% glucose (MMH) was tested at a pH of 6.4 ± 0.1, 7.1 ± 0.1, and 7.3 ± 0.1. Growth curves generated for F. noatunensis orientalis indicated that MMH at a pH of 6.4 ± 0.1 provided optimal growth. There were no significant differences in the growth curves obtained from isolates recovered from different fish species or from fresh or marine water. The pH of 6.4 ± 0.1 in the MMH media interfered with the inhibitory properties of the potentiated sulfonamides (ormetoprim-sulfadimethoxine and trimethoprim-sulfamethoxazole) when using the E. coli ATCC reference strain. Minimal inhibitory concentrations of eight antimicrobials (gentamicin, enrofloxacin, ampicillin, oxytetracycline, erythromycin, florfenicol, flumequine, and oxolinic acid) were similar for all F. noatunensis orientalis isolates. The in vitro susceptibility data provided here can provide a baseline for monitoring the development of antimicrobial resistance among F. noatunensis orientalis isolates, as well as provide valuable data in the development of potential therapeutics.

Received October 27, 2015; accepted April 13, 2016 Published online August 2, 2016     

Isolation and Partial Characterization of Extracellular Proteases Produced by Isolates of Flavobacterium columnare Derived from Channel Catfish

Abstract

Proteases of 23 isolates of Flavobacterium columnare derived primarily from channel catfish Ictalurus punctatus raised in the southeastern United States were isolated and partially characterized. The bacterial isolates were divided into two groups according to the apparent molecular masses of proteases after zymographic resolution by nonreducing, nondenaturing sodium dodccyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) with gelatin as the protease substrate. The 15 isolates in group 1 had two proteases with apparent molecular masses of 58 and 53.5 kilodaltons (kDa). Eight group-2 isolates produced three proteases with apparent molecular masses of 59.5, 48, and 44.5 kDa. Culture medium had an effect on the amount of protease produced by F. columnare LA 88–173. More protease was produced in a medium with low nutrients and salt (Ordal's medium) than in media with higher concentrations of nutrients or salts (TYES, Hsu-Shotts, modified Shieh's media). No differences were observed in the apparent molecular masses of the two proteases of F. columnare LA 88–173 produced in the various media or with different incubation times. Two proteases with apparent molecular masses of 58 and 53.5 kDa were seen as early as I d after inoculation, and these molecular masses did not change during the 7-d experiment. A sharp increase in protease production occurred during the first 24 h of incubation with minimal increase during the remaining 7 d of the experiment. All 23 isolates of F. columnare degraded the gelatin and casein incorporated into TYES agar medium but only 7 of the 23 isolates degraded elastin.     

Improved pulsed-field gel electrophoresis procedure for the analysis of Flavobacterium columnare isolates previously affected by DNA degradation

Flavobacterium columnare is a fresh water bacterium that causes columnaris diseases in over 36 fish species. Intra-species typing of F. columnare can be performed by pulsed-field gel electrophoresis (PFGE). However, this method is hampered by the degradation of chromosomal DNA in about 10% of strains. In the current study, DNA degradation problems caused by extracellular DNases were overcome by fixation of cells with formaldehyde prior to isolation. The results substantiate that after problems due to DNases are overcome, PFGE analysis is a reproducible highly discriminating epidemiological method for studying F. columnare isolates regardless of fish host.     

Technique for Identifying Flavobacterium columnare Using Whole-Cell Fatty Acid Profiles

Abstract

Isolates of Flavobacterium columnare (29 from diseased fish and three American Type Culture Collection cultures [ATCC 23463, 49512, 43622]) were identified by use of biochemical characteristics prior to generating whole-cell fatty acid profiles. The microbial identification system (MIS; Microbial ID, Newark, Delaware), a gas chromatography system, was used to generate the fatty acid profiles of F. columnare. The MIS contains databases of clinically and environmentally important bacteria that are represented by over 100 genera, including Flavobacterium spp. (F. aquatile [ATCC 11947] and F. mizutaii). Flavobacterium columnare is not included in the databases because it does not grow on standard media. Fatty acid profiles of F. columnare were generated with the CLIN40 protocol established by MIS after growth of the bacteria in modified Shieh broth. The fatty acid composition of F. columnare isolates determined by the CLIN40 method consisted of 10 major fatty acids (those present at levels > 1%): 11-methyl-dodecanoic acid (13:0 iso [the term “iso” designates the methyl group at the penultimate carbon atom]), 13-methyl tetradecenoic acid isomer G (15:1 iso G), 13-methyl tetradecanoic acid (15:0 iso), 12-methyl tetradecanoic acid (15:0 anteiso [the term “anteiso” designates the methyl group at the third carbon atom from the end]), pentadecanoic acid (15:0), 14-methyl pentadecanoic acid (16:0 iso), 3-hydroxy-13-methyl tetradecanoic acid (15:0 iso 3OH), 15-methyl cis-9-hexadecenoic acid (iso 17:1 ω9c), 3-hydroxy-14-methyl pentadecanoic acid (16:0 iso 3OH), and 3-hydroxy-15-methyl hexadecanoic acid (17:0 iso 3OH) (94.8% of profile). Five fatty acids found in the highest percentages from all isolates (CLIN40 method) included 15:1 iso G (16.12%), 15:0 iso (46.54%), 15:0 iso 3OH (6.81%), iso 17:1 ω9c (7.32%), and 17:0 iso 3OH (9.42%). Fatty acid profiles were also established by means of the MIS rapid protocol (RCLN50) in which identifications can be completed in 7 min instead of 20 min. Fatty acids found in the highest percentages for the RCLN50 protocol included 15:1 iso G (15.36%), 15:0 iso (43.03%), 15:0 iso 3OH (7.43%), iso 17:1 ω9c (6.83%), and 17:0 iso 3OH (10.17%). Both methods will allow reliable identification of F. columnare.     

Colorimetric Method of Loop-Mediated Isothermal Amplification with the Pre-Addition of Calcein for Detecting Flavobacterium columnare and its Assessment in Tilapia Farms

Flavobacterium columnare, the causative agent of columnaris disease in fish, affects many economically important freshwater fish species. A colorimetric method of loop-mediated isothermal amplification with the pre-addition of calcein (LAMP–calcein) was developed and used to detect the presence of F. columnare in farmed tilapia (Nile Tilapia Oreochromis niloticus and red tilapia [Nile Tilapia × Mozambique Tilapia O. mossambicus]) and rearing water. The detection method, based on a change in color from orange to green, could be performed within 45 min at 63°C. The method was highly specific, as it had no cross-detections with 14 other bacterial species, including other fish pathogens and two Flavobacterium species. The method has a minimum detection limit of 2.2 × 10² F. columnare CFU; thus, it is about 10 times more sensitive than conventional PCR. With this method, F. columnare was detected in gonad, gill, and blood samples from apparently healthy tilapia broodstock as well as in samples of fertilized eggs, newly hatched fry, and rearing water. The bacteria isolated from the blood were further characterized biochemically and found to be phenotypically identical to F. columnare. The amplified products from the LAMP–calcein method had 97% homology with the DNA sequence of F. columnare.

Received May 21, 2014; accepted August 10, 2014     

Characterization of four Flavobacterium columnare (Flexibacter columnaris) strains isolated from tropical fish

Four Flavobacterium columnare strains (AJS 1–4) were isolated from black mollies (Poecilia sphenops) and platies (Xiphophorus maculatus), showing white spots on the back, head and skin ulcers. The isolates developed characteristic rhizoid yellow pigmented colonies on Shieh agar and typical growth in Shieh broth. They were Gram-negative, filamentous bacteria exhibiting flexing movements. When compared to F. columnare strains isolated from temperate fish, it was noted that the four strains originating from tropical aquarium fish are more capable of growing at higher temperatures, the opposite being true for the strains isolated from temperate fish. Biochemical characterization and agglutination tests proved that the isolated strains could be classified as F. columnare. Low minimal inhibitory concentration (MIC) values were found for chloramphenicol, erythromycin, furazolidone, kanamycin, lincomycin, nalidixic acid, oxytetracycline and streptomycin. MIC values were high for colistin, sulfamethoxazole and neomycin. Pathogenicity studies were performed on black mollies. When these animals were submersed in an infective solution of the F. columnare strains, a marked difference in virulence was noted among the four isolated strains, strain AJS 1 being the most virulent one and strain AJS 4 being of low virulence.     

乳酸杆菌属对抗生素的耐药性

以分离自不同地区的发酵乳制品的44株乳杆菌为研究对象,通过对其进行16S rRNA的基因序列分析,鉴定为干酪乳杆菌、植物乳杆菌、发酵乳杆菌、瑞士乳杆菌和保加利亚乳杆菌.同时,研究了这5种乳杆菌的抗生素耐药性.结果显示,乳杆菌对不同的抗生素表现出不同的耐药性.     

湖南省部分规模猪场保育仔猪细菌感染调查及药敏检测分析

为调查规模猪场保育仔猪的细菌感染情况,文章对湖南常德、怀化、郴州等地10个规模猪场80头病、死仔猪进行细菌分离和药敏试验。结果显示:共在70头仔猪病料中分离到细菌,分离率为87.5%(70/80),其中大肠杆菌、猪链球菌、副猪嗜血杆菌、巴氏杆菌感染分别占52.5%(42/80)、48.8%(39/80)、30.0%(24/80)、15.0%(12/80),两种或两种以上细菌混合感染39头,占48.8%(39/80)。通过对分离菌株进行药敏试验,猪链球菌、副猪嗜血杆菌、巴氏杆菌对头孢曲松、多西环素、氟苯尼考、恩诺沙星较为敏感,而大肠杆菌则耐药严重,部分菌株对常用药物均耐药。可见规模猪场保育仔猪细菌混合感染情况较多,且耐药性严重的大肠杆菌为常见耐药菌之一。猪场应针对具体细菌感染情况采取针对性的用药方案,方可达到用药目的。该调查为湖南规模猪场保育仔猪细菌性疾病防控及指导用药提供了参考。     

蜂胶液中总黄酮含量测定方法的改进

研究改进后配置的芦丁对照品溶液对实验结果的影响.使用紫外一可见光分光光度法.分别以两种方法配制的对照品溶液对同一批号蜂胶液进行检测.得出20组数据,并进行假设检验,两种方法检测结果无显著性差异.说明改进后的标准品溶液能完全适用于蜂胶液总黄酮的测定.     

The effect of high total ammonia concentration on the survival of channel catfish experimentally infected with Flavobacterium columnare

Ammonia concentrations in water can affect the severity of Flavobacterium columnare infections in fish. Two trials lasting 7 d each were conducted to determine the effect of a single immersion flush treatment of total ammonia nitrogen (TAN; 15 mg/L) on the survival of channel catfish Ictalurus punctatus infected with E columnare; the chemical was added while the water flowed continuously through the tanks. Both trials consisted of four treatments: (1) no ammonia exposure and no bacterial challenge (control), (2) ammonia exposure only, (3) bacterial challenge only, and (4) both ammonia exposure and bacterial challenge. Two hours after exposure to ammonia, the highest un-ionized ammonia level was 0.43 mg/L. The percent un-ionized ammonia is based on TAN, temperature, and pH. Caudal fins from three fish in each treatment were sampled at 24 h posttreatment to be analyzed by quantitative real-time polymerase chain reaction (qPCR). No significant difference in survival (mean +/- SE) was noted between the channel catfish in treatment 1 (95.2 +/- 1.2%) and those in treatment 2 (95.6 +/- 1.0%); however, survival in both treatments 1 and 2 differed significantly from that in treatments 3 (8.5 + 4.5%) and 4 (41.8 +/- 12.7%). Treatment 4 catfish had significantly higher survival than treatment 3 catfish. Quantitative PCR data showed that treatment 4 fish had significantly less F. columnare (7.6 x 10(5)) than did treatment 3 fish (1.2 x 10(7)), and treatment 2 fish (8.5 x 10(3)) had significantly less bacteria than did treatment 1 fish (6.9 x 10(4)), indicating that ammonia limited the F. columnare infection. The highest mean concentration of the bacteria (3.9 x 10(7)) was found on moribund fish. The ammonia concentrations tested did not negatively influence fish survival but interfered with the infection process. An in vitro assay was also conducted to evaluate the direct effects of ammonia on F columnare.     

The Effect of High Total Ammonia Concentration on the Survival of Channel Catfish Experimentally Infected with Flavobacterium columnare

Abstract

Ammonia concentrations in water can affect the severity of Flavobacterium columnare infections in fish. Two trials lasting 7 d each were conducted to determine the effect of a single immersion flush treatment of total ammonia nitrogen (TAN; 15 mg/L) on the survival of channel catfish Ictalurus punctatus infected with F. columnare; the chemical was added while the water flowed continuously through the tanks. Both trials consisted of four treatments: (1) no ammonia exposure and no bacterial challenge (control), (2) ammonia exposure only, (3) bacterial challenge only, and (4) both ammonia exposure and bacterial challenge. Two hours after exposure to ammonia, the highest un-ionized ammonia level was 0.43 mg/L. The percent un-ionized ammonia is based on TAN, temperature, and pH. Caudal fins from three fish in each treatment were sampled at 24 h posttreatment to be analyzed by quantitative real-time polymerase chain reaction (qPCR). No significant difference in survival (mean ± SE) was noted between the channel catfish in treatment 1 (95.2 ± 1.2%) and those in treatment 2 (95.6 ± 1.0%); however, survival in both treatments 1 and 2 differed significantly from that in treatments 3 (8.5 ± 4.5%) and 4 (41.8 ± 12.7%). Treatment 4 catfish had significantly higher survival than treatment 3 catfish. Quantitative PCR data showed that treatment 4 fish had significantly less F. columnare (7.6 × 10⁵) than did treatment 3 fish (1.2 × 10⁷), and treatment 2 fish (8.5 × 10³) had significantly less bacteria than did treatment 1 fish (6.9 × 10⁴), indicating that ammonia limited the F. columnare infection. The highest mean concentration of the bacteria (3.9 × 10⁷) was found on moribund fish. The ammonia concentrations tested did not negatively influence fish survival but interfered with the infection process. An in vitro assay was also conducted to evaluate the direct effects of ammonia on F. columnare.

Received September 15, 2010; accepted May 7, 2011     

Determination of Phylogenetic Relationships of Flavobacterium psychrophilum (Flexibacter psychrophilus), Flavobacterium columnare (Flexibacter columnaris), and Flexibacter maritimus by Sequence Analysis of 16S Ribosomal RNA Genes Amplified by Polymerase Chain Reaction

Abstract

The nearly complete small subunit 16S ribosomal RNA (rRNA) gene sequence amplified by polymerase chain reaction was determined for Flavobacterium psychrophilum (formerly Flexibacter psychrophilus) by using automated nucleotide sequencing. The sequence was found to be 1,465 base pairs (bp) in length, a size consistent with previously determined sequences for 14 other bacterial species from various taxa, including the yellow-pigmented bacteria. Sequence signatures confirmed that this organism was a member of the bacterial division Bacteroides–Flavobacterium. Parsimonious and additive phylogenetic trees were constructed with homology, and pairwise evolutionary distances were used to estimate phylogenetic relationships. Data show that F. psychrophilum, F. columnare, and Flexibacter maritimus are closely related, have a common descent, and represent a distinct group within the division Bacteroides–Flavobacterium. This group also included other organisms from the genera Flavobacterium and Cytophaga. Further, Flavobacterium aquatile, the type species for the genus Flavobacterium, was also determined to be a member of this “Flavobacterium–Cytophaga–Flexibacter subcomplex.” This supports previous assertions that the type strain of Flavobacterium should be changed to a more representative species such as Flavobacterium breve.     

Antagonistic effect of indigenous skin bacteria of brook charr (Salvelinus fontinalis) against Flavobacterium columnare and F. psychrophilum

Industrial fish production exposes fish to potentially stressful conditions, which in turn may induce infections by opportunistic pathogens. Probiotics appear to be a promising way to prevent opportunistic infections in aquaculture. In this study, we tested the inhibitory potential of endogenous bacterial communities found in the mucus of brook charr (Salvelinus fontinalis) against two major pathogens Flavobacterium columnare and Flavobacterium psychrophilum. Nine bacterial strains were isolated from brook charr skin mucus and tested for potential antagonistic activity. Results from both agar diffusion assays and broth co-culture assays showed the presence of antagonism. We identified seven bacterial strains, collected from unstressed fish, which exerted strong antagonism against F. psychrophilum and/or F. columnare. These strains were mixed and used to treat columnaris disease in an in vivo experiment in which four distinct fish families were tested. This treatment resulted in a decrease of mortality (54-86%) across fish families indicating that candidates from the host microbiota are potentially suitable for probiotic development. This would allow for the efficient (ability to adhere and colonize the host mucus) and durable management (antagonistic effect against pathogens which would be harmless for the host and safe for its environment) of opportunistic diseases in aquaculture.     

The association of Flavobacterium columnare strains of high and low virulence with gill tissue of black mollies (Poecilia sphenops).

The ability of a high virulence strain (AJS 1) and a low virulence strain (AJS 4) of Flavobacterium columnare (Flexibacter columnaris) to attach to the gills of black mollies (Poecilia sphenops) was investigated. For that purpose, two groups of 25 black mollies each were immersed in a bath containing 10(6) CFU/ml of F. columnare AJS 1 or AJS 4. At regular intervals from 1 to 12 h after the contact infection, fish were sacrificed and gills, skin, spleen and heart were sampled for bacteriology. Samples of the gills were taken for immunohistochemical and electron microscopic examination. Bacteriological examination proved that the number of gill-associated F. columnare was higher for AJS 1 than for AJS 4. Strain AJS 1 was isolated from the heart and spleen of 6 and 1 of the 16 examined animals, respectively. Strain AJS 4 was not isolated from the internal organs of any fish. When examined immunohistochemically, strain AJS 1 was found closely associated with gill epithelium whereas this was not the case for strain AJS 4. The adherence of bacteria to the gill tissue challenged with the virulent strain AJS 1 was also clearly demonstrated using scanning and transmission electron microscopy. These results indicate that adhesion of F. columnare to the gill tissue constitutes an important step in pathogenesis.     

大通量的抗生素药敏检测盒的设计及应用

基于琼脂稀释法,设计制作了96点阵抗生素药敏检测盒。以NCCLS（2007）抗生素最低抑菌浓度(minimal inhibitory concentration,MIC)微量稀释法为对照,测定了分离自规模化猪场饲养员和幼儿园、中学学生的98株大肠杆菌对恩诺沙星、环丙沙星、诺氟沙星和中药三黄汤的药物敏感性。结果表明,两种方法测定的最低杀菌浓度(minimal bactericidal concentration,MBC)、MIC都表现高度对称,其差值的平均值都在一倍稀释度内,在差值0.5 μg/mL范围内都达到90%以上的符合。MBC与MIC高度相关,r=0.94~0.99。这些指标说明两种方法测定的MBC与MIC平行性好。以总体的抗生素敏感性频率比较,两种方法测定值也很相近,符合率为93.93%。本检测盒还可以准确测定中药三黄汤对大肠杆菌的MIC（125～500 mg/mL）。96点阵抗生素药敏检测盒是适合于实验室进行大规模抗药性监测的新工具,具有简易、准确、快速、大通量、节约的优点。配合96孔培养板,一次可以测定96株病原菌对一种抗生素或中药的敏感性。     

猪胸膜肺炎放线杆菌四川株的分离鉴定及药物敏感性试验

从四川某猪场疑似胸膜肺炎放线杆菌感染的病料中分离到一株革兰氏阴性小球杆菌。经染色镜检、生化试验、PCR检测、血清型检测,致病性试验和药敏试验,鉴定该菌为猪胸膜肺炎放线杆菌1型,且具有较强致病性。药敏试验结果表明,该菌对四环素、甲氧苄啶,以及链霉素、庆大霉素、卡那霉素、丁胺卡那、新霉素等氨基糖苷类药物高度耐药,对氯霉素、氟苯尼考、氨苄西林、阿莫西林、羧苄西林和恩诺沙星、诺氟沙星等喹诺酮类受试药物敏感。     

Antimicrobial Susceptibility Patterns of Clostridium difficile Isolates from Family Dairy Farms

A significant risk factor for developing Clostridium difficile infection (CDI) in humans and animals is associated with the antimicrobial use. It has often been hypothesized that farm animals could be the source for human infection with Clostridium difficile (CD). In the European Union, family‐run dairy farms are the predominant farming model, which are more interlinked within the community compared to large‐scale intensive dairy or beef farms. Therefore, it is important to investigate antimicrobial susceptibility patterns of CD in such environment. A total of 159 CD isolates from 20 family dairy farms were tested with a customized broth microdilution plate for their antimicrobial resistance. Seventeen antimicrobials were selected (amoxicillin, ceftriaxone, clindamycin, daptomycin, erythromycin, fusidic acid, imipenem, levofloxacin, linezolid, metronidazole, moxifloxacin, oxacillin, rifampicin, tetracycline, tigecycline, trimethoprim/sulfamethoxazole and vancomycin), which are commonly used for treatment of CDI in veterinary and human medicine, or were previously applied in CD epidemiological studies. Antimicrobials, which are used for treatment of CDI in humans (metronidazole, vancomycin, fusidic acid, tigecycline, linezolid) inhibited CD growth in vitro. Most CD isolates were resistant to erythromycin (93.1%), daptomycin (69.2%) and clindamycin (46.5%). High multiple‐resistance was found in CD ribotype 012 (n = 5, 100%), some CD SLO 060 (n = 4, 25%) and one CD 033 (n = 1, 1.1%). High multiple‐resistance in this study was linked with CD ribotypes and not with the origin of CD. The low prevalence of these ribotypes (6.3%; 10/159) indicates that family‐run dairy farms are an unlikely source of CD with multiple‐resistance to antimicrobials.     

副猪嗜血杆菌的分离鉴定及药敏试验

黑龙江省近几年频繁发生以多发性浆膜炎、关节炎、脑膜炎以及急性死亡为特征的传染病,为猪场造成严重经济损失,为了确诊是否有副猪嗜血杆菌感染,采集病死猪肝脏、脾脏和肺脏等病料,进行副猪嗜血杆菌的分离;对其理化特性进行鉴定,应用PCR方法对其16S rRNA基因扩增后进行克隆测序,将测序结果在GenBank上进行BLAST分析,把相近的基因序列应用DNAStar软件进行同源性和进化关系分析;用分离菌株进行药物敏感性试验,筛选敏感药物。结果分离出一种具有多形性的NAD依赖性菌株,经鉴定为副猪嗜血杆菌;16S rRNA基因进化分析结果表明,分离菌株与以往报道的副猪嗜血杆菌中国、日本和美国分离株属于同一亚群,而西班牙分离株属于单独的亚群;分离菌株对壮观霉素和头孢拉定高度敏感。     

硫酸粘杆菌素微生物检定法的改进

本文以大肠杆菌标准菌株(Escherichia coli CMCC(B)44103)为检测菌,对硫酸粘杆菌素的微生物检测方法进行了研究.研究结果表明,本方法检测限为0.2μg/ml,与国外所用的以博德特氏菌ATCC4617为检测菌的检测限基本一致,其质量控制样品的日内、日间变异系数均小于10%,平均回收率94%～100.8%,并且操作简便,是一种较好的替代方法,可用于硫酸粘杆菌素的药代动力学研究或进行残留检测.