The microbial environment of the ear canal in health and disease

Otitis externa is an important disease of dogs and, to a lesser degree, cats. The yeast Malassezia canis is the most common organism demonstrated in ear infections. Coagulase-positive staphylococci are the most common bacteria isolated, often occurring with M. canis. Pseudomonas aeruginosa and Proteus are gram-negative bacteria that are frequently encountered. Other bacteria, such as beta-hemolytic streptococci, enterococci, Escherichia coli, and Corynebacterium are also important. Malassezia canis, coagulase-positive staphylococci, and Pseudomonas aeruginosa commonly occur as monoinfections, whereas other bacteria are typically associated with mixed infections. Smears may provide rapid diagnosis on which to base empiric treatment. Cultures should be considered in recurrent or refractory cases, especially those involving gram-negative bacteria. In practices that are not equipped to perform culture and susceptibility tests, it is advisable to seek the advice of a microbiology laboratory.     

The role of Pityrosporum pachydermatis in otitis externa of dogs: evaluation of a treatment with miconazole.

The bacterial and mycotic flora were assessed in 158 ears of dogs with otitis externa and in 101 ears of healthy control dogs. Pityrosporum pachydermatis occurred in 57 per cent of ears with otitis externa and in 17 per cent of clinically healthy ears. Staphylococci and Pseudomonas aeruginosa were the predominant bacteria in otitic ears, micrococci and Bacillus spp were the most frequent isolates from clinically healthy ears. P pachydermatis, Ps aeruginosa and Candida tropicalis occurred in monoculture in a significant number of mainly chronic cases of otitis externa. A combination preparation, containing miconazole, polymyzin B and prednisolone, was highly effective in controlling the clinical signs of otitis externa and eliminating flora from the affected ears. The data presented suggest that yeasts, and especially P pachydermatis, may be significant pathogens in otitis externa and that antimycotic treatment is an essential part of the treatment of otitis externa in dogs.     

Antibiotic susceptibility of bacteria isolated from cats with ulcerative keratitis in Taiwan

Objectives: To assess the antibiotic susceptibility of bacteria isolated from corneal ulcers in cats. Methods: A total of 92 cats with infected corneal ulcers were swabbed for bacterial culture and the antibiotic sensitivity of the isolates analysed. Results: Bacteria were isolated from 54 of 92 infected eyes with corneal ulcers and purulent discharge. A total of 59 bacterial isolates were obtained from the 54 ulcers. The ratio of Gram-positive to Gram-negative isolates was approximately 3:1. The most commonly isolated Gram-positive bacteria were Staphylococcus species (51 per cent of all isolates), while Pseudomonas aeruginosa (13.5 per cent of all isolates) was the most common Gram-negative bacteria isolated. The Gram-negative isolates demonstrated a greater incidence of antibiotic resistance than the Gram-positive ones. The most effective antibiotics against the isolates were ciprofloxacin, tobramycin and gentamicin, with erythromycin and lincomycin showing the greatest number of resistant isolates. Clinical Significance: Staphylococcus and Pseudomonas species were the most common Gram-positive and Gram-negative bacteria, respectively, isolated from feline eyes with ulcerative keratitis. The second-generation fluoroquinolone, ciprofloxacin and the aminoglycoside gentamicin were found to be highly effective against the majority of isolates.     

竹鼠绿脓杆菌的分离和鉴定

对广西某养殖户送检的病竹鼠进行病理剖检及细菌分离鉴定,根据发病临床症状,病理变化,病原分离鉴定及致病性试验结果,确诊为绿脓杆菌感染。对分离菌进行药敏试验,结果显示所分离到的绿脓杆菌细菌对氧氟沙星、诺氟沙星、环丙沙星、庆大霉素、阿米卡星和阿奇霉素敏感;对头孢他啶、头孢西丁、利福平和阿莫西林耐药。     

Antibiotic susceptibility of bacterial isolates from corneal ulcers of dogs in Taiwan

OBJECTIVES: To analyse antibiotic susceptibility of bacteria associated with ulcerative keratitis in dogs. METHODS: Bacteria isolated from 190 eyes with ulcerative keratitis were identified, and the antibiotic susceptibility of isolates was studied. RESULTS: In total, 258 species of bacteria were isolated from the 190 eyes. Of the isolates, 78 per cent were Gram-positive and 28 per cent were Gram-negative bacteria. The most commonly isolated Gram-positive bacteria in dogs were Staphylococcus spp (49 per cent), Streptococcus spp (7 per cent) and Corynebacterium spp (7 per cent); while Pseudomonas aeruginosa (7.6 per cent) and Escherichia coli (5.8 per cent) were the commonest Gram-negative pathogens. Resistance to commonly used ophthalmic antibiotics was seen in Staphylococcus, Streptococcus, Corynebacterium, Pseudomonas and Escherichia species isolates. CLINICAL SIGNIFICANCE: Isolates from dogs with corneal ulcers in Taiwan may be resistant to several commonly used ophthalmic preparations. Ciprofloxacin showed good action against most isolates, with the notable exception of Streptococcus species. Chloramphenicol or cephalothin had the best in vitro action against the Streptococcus species isolates.     

The bovine innate immune response during experimentally-induced Pseudomonas aeruginosa mastitis

Almost half of all clinical cases of mastitis are caused by Gram-negative bacteria. Among these bacteria, intramammary infection by Pseudomonas aeruginosa remains one of the most refractory to antibiotic therapy. The ability to recognize potentially harmful pathogens whether previously encountered or not, as well as the induction of an initial pro-inflammatory response to these pathogens, are critical components of host innate immunity. Although the innate immune response to another Gram-negative mastitis-causing pathogen, Escherichia coli, has been well-characterized, little is known about the response to other Gram-negative bacteria, including P. aeruginosa. The objective of the current study was to characterize the systemic and localized bovine innate immune response to intramammary infection with P. aeruginosa. The contralateral quarters of ten mid-lactating Holstein cows were challenged with either saline or P. aeruginosa. Following the establishment of infection, milk samples were collected and assayed for changes in cytokine and growth factor concentrations, complement activation, and changes in the levels of soluble CD14 (sCD14) and lipopolysaccharide (LPS)-binding protein (LBP), two accessory molecules involved in host recognition of Gram-negative bacteria. Initial increases in milk somatic cell counts were evident within 12h of experimental challenge and remained elevated for >or=3 weeks. Increased permeability of the mammary gland vasculature, as evidenced by elevated milk levels of BSA, was initially observed 20 h post-infection and persisted for 2 weeks. Within 32 h of challenge, increased levels of IL-8, TNF-alpha, IL-10, and IL-12 were detected, however, the elevated levels of these cytokines were not sustained for longer than a 24h period. In contrast, elevations in IL-1beta, IFN-gamma, TGF-alpha, TGF-beta1, TGF-beta2, sCD14, LBP, and activated complement factor 5 (C5a) were sustained for periods of >48 h. Systemic changes were characterized by elevated body temperature, induction of the acute phase protein synthesis of serum amyloid A and LBP, and a transient decrease in circulating neutrophils and lymphocytes. Together, these data demonstrate the capability of the mammary gland to mount a robust innate immune response to P. aeruginosa that is characterized by the induction of pro-inflammatory cytokines, complement activation, and increased levels of accessory molecules involved in Gram-negative bacterial recognition.     

检测水貂出血性肺炎绿脓杆菌的比色LAMP法的建立与应用

为了快速有效检测水貂出血性肺炎病原绿脓杆菌,本研究结合金属指示剂HNB特性建立了快速检测绿脓杆菌比色LAMP法。根据绿脓杆菌外毒素A基因设计引物,建立了检测LAMP法;并且对该方法进行了特异性、灵敏性分析;同时进行了初步应用。结果显示,该方法特异性强,灵敏性好,可检测167CFU/mL的菌体,对病貂肺脏检出率高。结果表明,该比色LAMP法可以有效地检测水貂出血性肺炎绿脓杆菌。     

Establishment and Preliminary Application of Triple PCR to Identify Three Kinds of Conditional Pathogenic Bacteria

The aim of this study was to establish a simultaneous triple PCR detection method for Staphylococcus aureus (S.aureus),Pseudomonas aeruginosa (P.aeruginosa) and Klebsiella pneumoniae (K.pneumoniae).Three pairs of specific primers had been designed according to nuc gene of S.aureus,toxR gene of P.aeruginosa and PhoE gene of K.pneumoniae.The triple PCR reaction conditions were optimized on the basis of single PCR methods.At the same time,specificity,sensitivity and repeatability tests of the triple PCR method were studied,and the results of bacteria isolation and culture and the triple PCR method were compared.The results showed that the amplification product sizes were 484,278 and 368 bp,respectively.The optimal annealing temperature was 56 to 59 ℃,the concentrations of primers were all 0.2 μmol/L,dNTP concentration was 200 μmol/L,Mg²⁺ concentration was 2.5 mmol/L.The specificity test showed that there was no cross reaction between these three bacteria templates and other eight kinds of common bacteria templates,such as Bordetella bronchiseptica.The minimum of simultaneous detection of three bacteria genomic DNA was 10^-5 ng/μL.The results of three repeatability tests of triple PCR were the same which indicated the repeatability was good.30 samples from mice were detected by bacteria isolation and culture and the triple PCR method,the detection rate of triple PCR was slightly higher than the bacteria isolation and culture.The positive samples detected by bacteria isolation and culture were also positive detected by triple PCR.The results showed that a specific,sensitive and efficient triple PCR system had been established and could provide the technical support for bacteria detection and epidemiological investigation of laboratory animals.     

3种条件性致病菌三重PCR检测方法的建立及初步应用

本研究旨在建立一种能同时检测金黄色葡萄球菌(Staphylococcus aureus,S.aureus)、绿脓杆菌(Pseudomonas aeruginosa,P.aeruginosa)、肺炎克雷伯杆菌(Klebsiella pneumoniae,K.pneumoniae)的三重PCR检测方法.根据金黄色葡萄球菌nuc基因、绿脓杆菌toxR基因、肺炎克雷伯杆菌PhoE基因设计并合成引物,在单一PCR条件基础上优化建立三重PCR反应条件,并进行特异性、敏感性和重复性分析及与细菌分离培养的比对试验.结果显示,3对引物均能特异性扩增出目的条带,大小分别为484、278和368 bp.最佳退火温度在56~59 ℃之间,引物浓度均为0.2 μmol/L,dNTP浓度为200 μmol/L,Mg²⁺浓度为2.5 mmol/L.3种细菌间无交叉反应.对支气管鲍特杆菌等其他8种实验动物常见致病菌均无交叉反应.最低能同时检测到10^-5 ng/μL的细菌基因组DNA.3次重复结果一致,表明建立的三重PCR方法重复性好.同时采用细菌分离培养法和三重PCR方法对30份实验小鼠样本进行检测,对比结果显示三重PCR方法检出率略高于细菌分离培养法,细菌分离培养法呈阳性的样品,三重PCR方法均能检出.结果表明,本试验建立的三重PCR检测方法具有特异、敏感、高效等优点,为实验动物细菌快速检测和流行病学调查提供了技术支持.     

Research Progress on Biofilms in Pseudomonas aeruginosa

The opportunistic pathogen Pseudomonas aeruginosa is a gram-negative bacteria which can form biofilms.In this review,we summarized the biology mechanism of biofilms in Pseudomonas aeruginosa,including Pseudomonas aeruginosa adhesion,extracellular polysaccharide Psl and Pel,alginate participated in the biofilm maturation,and quorum sensing systems regulated the formation of biofilm in Pseudomonas aeruginosa,and the drug target on biofilm.The bacterial biofilms protected the bacteria from unsuitable environmental conditions.Through researching the structure and the pathogenesis of Pseudomonas aeruginosa,and its resistance molecular mechanisms,we could adjust or regulate the expression of biofilm-associated factors,optimize the anti-infection treatment in Pseudomonas aeruginosa.     

铜绿假单胞菌生物膜研究进展

条件致病菌铜绿假单胞菌（PA）是一种能形成生物膜的革兰氏阴性菌,作者综述了PA生物膜形成的生物学机制,包括菌体黏附、胞外多糖Psl和Pel、藻朊酸盐等参与细菌生物膜成熟的过程及群体感应系统调节相关因子表达,从而调控细菌形成生物膜应对不良环境。此外还概括了将生物膜作为靶点开发的药物等生物膜相关的研究进展。生物膜是菌体逃避有害刺激的护盾,研究其结构、形成及致病机理,了解PA产生耐药性的分子机制,对于通过调节生物膜形成或调控生物膜相关因子的表达进而优化PA的抗感染治疗有十分重要的意义。     

竹鼠绿脓杆菌的分离鉴定及耐药性分析

对广西某竹鼠养殖户送检的病竹鼠进行病理剖检及细菌分离鉴定,据临床发病症状、病理变化、病原分离鉴定及致病性试验结果,确诊为绿脓杆菌感染。动物致病力试验结果表明该分离株具有较强的致病力;对分离菌进行药敏试验,结果显示所分离到的绿脓杆菌对氧氟沙星、诺氟沙星、环丙沙星、庆大霉素、阿米卡星和阿奇霉素敏感,对头孢他啶、头孢西丁、利福平和阿莫西林耐药。     

Use of specific sugars to inhibit bacterial adherence to equine endometrium in vitro

OBJECTIVE: To determine whether specific sugars inhibit adhesion of Streptococcus zooepidemicus, Pseudomonas aeruginosa, and Escherichia coli to equine endometrial epithelial cells in vitro. SAMPLE POPULATION: Endometrial biopsy specimens collected during estrus from 7 healthy mares. PROCEDURE: Endometrial specimens on glass slides were incubated for 30 minutes at 4 C with suspensions of S. zooepidemicus, P. aeruginosa, or E. coli in phosphate-buffered saline solution (PBSS) alone or with various concentrations of D-(+)-mannose, N-acetyl-D-glucosamine, N-acetyl-D-galactosamine, D-(+)-glucose, galactose, or N-acetyl-neuraminic acid. Inhibition of bacterial adherence was determined by comparing adhesion of bacteria (i.e., percentage of glandular epithelial cells with adherent bacteria) suspended in each sugar solution with that of bacteria suspended in PBSS. RESULTS: Mannose and N-acetyl-D-galactosamine inhibited adhesion of E. coli and P. aeruginosa to epithelial cells, whereas only mannose inhibited adhesion of S. zooepidemicus. The other sugars did not affect bacterial adherence. CONCLUSIONS AND CLINICAL RELEVANCE: Mannose and N-acetyl-D-galactosamine appear to play a role in adhesion of S. zooepidemicus, P. aeruginosa, and E. coli to equine endometrium. In horses with uterine infections, use of sugars to competitively displace bacteria from attachment sites on cells may provide an adjunct to antibiotic treatment.     

绿脓杆菌研究进展

绿脓杆菌存在于环境和正常畜禽体内,是一种条件致病菌。近年来,随着动物疾病研究的深入,有关绿脓杆菌感染的报道也越来越多。从病原学、流行病学、发病机理、临床学等方面对绿脓杆菌的研究进展进行了综述,以期为控制绿脓杆菌的感染奠定基础。     

急性临床型奶牛乳房炎病原菌的分离鉴定与药敏试验

本文对一起群发性急性临床型奶牛乳房炎进行了病原分离鉴定、致病性及药敏试验,结果表明该乳房炎是由金黄色葡萄球菌、停乳链球菌和铜绿假单胞菌混合感染引起,其中金黄色葡萄球菌和铜绿假单胞菌对小鼠表现出较强的致病性,三株菌对兽医临床常用抗生素均表现出较强的耐药性。     

一例雏鸡铜绿假单胞菌的分离鉴定及其16SrRNA基因序列分析

为对死亡雏鸡进行病因诊断,通过大体剖检、细菌分离、生化鉴定,证实为铜绿假单胞菌感染。测定了该分离株的16SrRNA基因序列,并与GenBank中收录的序列比较,结果发现所分离的铜绿假单胞茵及参考株的16SrDNA基因序列极其保守,相似性达99％～100％;与大肠杆菌、沙门氏菌、巴氏杆菌的相似性差异约为9％;与鸭疫里默氏...     

Relationship between the immune response of sheep and the population dynamics of bacteria isolated from fleecerot lesions.

In sheep wetted by rain, proliferation of bacteria in the skin-fleece microenvironment invariably discolours the fleece and causes a dermatitic condition known as fleecerot. The changes in population dynamics of fleece bacteria were analysed by carrying out skin washings at randomly selected sites on the back of sheep before, and at 48 h and 96 h after exposure to rain. Gram-positive rods belonging to Bacillus species (10(2)-10(4) cfu/cm2) predominated in dry fleece. Gram-positive cocci (e.g. Micrococcus and Staphylococcus species) as well as Gram-negative rods (pseudomonads) were also present but in lower abundance (less than 10(2) cfu/cm2). Fleece bacterial populations generally increased in numbers during the first 24-48 h of wetting. By 96 h however, skin washings showed a preponderance of Pseudomonas aeruginosa (10(4)-10(6) cfu/cm2) and to a lesser extent, pigmented Micrococcus species. Growth of fleece bacteria was associated with a characteristic green or yellow/orange staining of fleece. Fewer species of bacteria were isolated from sheep showing green staining while those animals with yellow/orange discolourations appeared to have a more mixed microflora composition. The predominance of P. aeruginosa in the wet fleece of sheep displaying either green or yellow/orange bacterial stain, was accompanied by a significant serological response against this species. Since skin bacteria have never been observed to penetrate cutaneously in skin sections biopsied from fleecerot sites, it must be concluded that the sheep skin is sensitized by continuous exposure to antigens that are associated with or released by P. aeruginosa.     

Bacteriology and mycology of otitis externa in dogs]

The bacterial and fungal flora of 1118 ears of dogs with otitis externa and 100 ears of healthy control dogs were studied in order to isolate the causative agents. The yeast Malassezia pachydermatis (56%) was by far the most common organism in otitic dogs followed by the bacteria Staphylococcus intermedius (23%), Pseudomonas aeruginosa (12%), Proteus spp. (6%) and Streptococcus canis (5%). A statistical analysis of observed results showed that the incidence of these organisms is significant in otitic dogs. Many strains of S.intermedius, P.aeruginosa and Proteus spp. are resistant to antimicrobial agents commonly used to treat otitis externa. Therefore an antimicrobial susceptibility testing was performed using "Cobas Bact" for these bacterias. Furthermore, 80 strains of M.pachydermatis were submitted to identification-kits (API 20 CAUX, API STAPH, Cobas Micro). The observed results showed that an identification with these tests was not possible.     

麝化脓病细菌性病原诊断鉴定

目的:寻找麝化脓病细菌性主要病原,为防治麝化脓病以及麝化脓病病原的研究奠定基础。方法:病料组织触片镜检,兰氏染色后镜检,用LB培养基、鲜血培养基对56份病料进行病原菌的分离培养,观察菌落形态,兰氏染色镜检,用微量生化反应管对细菌进行生化试验,对分离得到的菌株用纸片法进行药物敏感性试验。结果:从分离菌的形态、培养特征、生理生化特性等,可以证实分离的3种菌分别为巴斯德菌(Pasteurella spp)或绿脓杆菌(Pseudomonas aeruginosa)或耶尔森菌(Yersinia spp)、化脓隐秘杆菌(Actinomyces pyogenes)和葡萄球菌(Staphylo-coccus spp)。结论:根据流行病学特点、对死亡麝病变组织的剖解检查和病原菌分离培养鉴定的结果,可以证实引起麝发生化脓病病变的病原菌为巴斯德菌或绿脓杆菌或耶尔森菌、化脓隐秘杆菌和葡萄球菌。临床治疗首选药物为左氧氟沙星、磺胺、先锋霉素Ⅳ。     

致仔猪肠壁囊泡性脓肿铜绿假单胞菌的生物学特性研究

为研究猪肠壁囊泡性囊肿的可能病原,从发病猪肠道囊泡分离细菌,从11头发病死亡猪肠壁囊泡分离到1株共同性的细菌,对其进行了细菌生化鉴定和16sRNA测序,结果鉴定该株细菌为铜绿假单胞菌。通过药敏试验和小鼠攻毒试验,证实该株细菌具有多重耐药性和强致病性。本研究首次报道了铜绿假单胞菌致仔猪肠壁裳泡性脓肿的病例,并证明了假铜绿单胞菌强的致病作用,为深入铜绿假单胞菌对畜禽养殖的威胁及条件致病菌的变异奠定基础。