Structure of the microflora of suppurative wounds and its sensitivity to antibiotics]

Clinico-bacteriological examination of patients with purulent infections showed that Staphylococcus was the predominating microflora in the wounds. Simultaneously an increasing role of gram-negative conditionally pathogenic bacteria was shown. Multiple drug resistance was found in the organisms tested. The highest sensitivity levels were observed to gentamicin, kanamycin, tetracycline, levomycetin. It was shown by means of special typing methods that staphylococci of phage group III and Ps. Aeruginosa of serotype II predominated in the infected wounds. When the pathological material contained the antibiotic resistant cultures of Ps. aeruginosa, Proteus, Klebsiella and toxigenic strains of Staphylococcus, a tendency for prolongation of the suppurative process was observed.     

Effect of a controlled abacterial environment on the Pseudomonas aeruginosa infection of suppurative wounds

The present work deals with the data on the isolation rate of P. aeruginosa from suppurative wounds of different origin during their treatment by the commonly used methods under dressings and by the open method under the conditions of controlled germ-free environment. The results of the immunotyping of P. aeruginosa strains isolated from patients treated by different methods are presented. The dynamics of changes in the isolation rate of P. aeruginosa at different periods of treatment, both by the open method and with the use of dressings, is shown. Among P. aeruginosa strains isolated from suppurative wounds, those belonging to immunotypes 6, 7 and 2, as well as nontyping strains, occurred most frequently. Treatment in the controlled germ-free environment permits the protection of the wound surface from hospital infection. During treatment with the use of dressings the cases of hospital infection were revealed (31.3%). Such infection occurred, as a rule, at a later period of treatment.     

Epidemiological investigation and typing of Candida glabrata clinical isolates by FTIR spectroscopy

Candida glabrata has emerged as one of the leading agents of fungal infections and strain typing is essential for epidemiological investigation that is generally achieved by molecular techniques. In this work, we studied twenty-nine C. glabrata strains isolated from different patients, using a phenotypic approach based on Fourier Transform Infrared (FTIR) spectroscopy, which has been in a previous study successfully applied as a rapid typing method for Candida albicans. A two-step procedure was used for the analysis. The first step included sixteen strains for the internal validation phase, which aimed at finding the spectral windows that would provide the best differentiation between strains. In this phase, hierarchical cluster analysis (HCA) carried out using three spectral windows (900-1200, 1540-1800, 2800-3000 cm(-1)) allowed to obtain the best classification, where each patient strains could be clustered together. A genotypic technique based on randomly amplified polymorphic DNA-analysis (RAPD) confirmed these results. In a second step, the external validation phase, thirteen other clinical strains of C. glabrata isolated from multiple sites in four ICU patients, were tested by FTIR spectroscopy. The analysis was based on the spectral regions previously found in the first step. HCA classification of the strains gave four groups, one group per patient. These results suggest that no inter-human transmission took place. This study shows the potential of FTIR approach for typing of C. glabrata with several advantages compared to other techniques. FTIR typing is fast, effective, and reagent free. Moreover, it is applicable to all micro-organisms and requires a small quantity of biomass.     

RAPD typing of clinical isolates of Staphylococcus haemolyticus

The randomly amplified polymorphic DNA (RAPD) assay was used to generate DNA fingerprints from clinical isolates of Staphylococcus haemolyticus isolated from patients treated with continuous ambulatory peritoneal dialysis and previously subjected to a combination of typing methods. The RAPD profiles generated with one of six randomly designed 10-mer primers allowed visual discrimination of strains. Good correlation with the original typing scheme was achieved but RAPD typing allowed discrimination of strains previously indistinguishable.     

Epidemiological investigation and typing of Candida glabrata clinical isolates by FTIR spectroscopy

Candida glabrata has emerged as one of the leading agents of fungal infections and strain typing is essential for epidemiological investigation that is generally achieved by molecular techniques. In this work, we studied twenty-nine C. glabrata strains isolated from different patients, using a phenotypic approach based on Fourier Transform Infrared (FTIR) spectroscopy, which has been in a previous study successfully applied as a rapid typing method for Candida albicans. A two-step procedure was used for the analysis. The first step included sixteen strains for the internal validation phase, which aimed at finding the spectral windows that would provide the best differentiation between strains. In this phase, hierarchical cluster analysis (HCA) carried out using three spectral windows (900–1200, 1540–1800, 2800–3000 cm^− 1) allowed to obtain the best classification, where each patient strains could be clustered together. A genotypic technique based on randomly amplified polymorphic DNA-analysis (RAPD) confirmed these results. In a second step, the external validation phase, thirteen other clinical strains of C. glabrata isolated from multiple sites in four ICU patients, were tested by FTIR spectroscopy. The analysis was based on the spectral regions previously found in the first step. HCA classification of the strains gave four groups, one group per patient. These results suggest that no inter-human transmission took place. This study shows the potential of FTIR approach for typing of C. glabrata with several advantages compared to other techniques. FTIR typing is fast, effective, and reagent free. Moreover, it is applicable to all micro-organisms and requires a small quantity of biomass.     

Acute suppurative thyroiditis in appearance of unusual neck mass

We report on three adult cases of acute suppurative thyroiditis due to infection through a piriform sinus fistula. In these cases, the lesion did not exhibit the clinical symptoms characteristic of acute suppuration, but appeared as an unusual neck mass. Repeated barium meal roentgenograms, however, demonstrated the internal fistula in two of the three cases. Acute suppurative thyroiditis may not be so rare even in adults. When acute suppurative thyroiditis occurs in adults with scanty symptoms of acute suppuration, the piriform sinus fistula could easily go undetected. Recognition of acute suppurative thyroiditis in adults per se seems to be important for correct diagnosis.     

Genomic Characterization of a Large Outbreak of Legionella pneumophila Serogroup 1 Strains in Quebec City, 2012

During the summer of 2012, a major Legionella pneumophila serogroup 1 outbreak occurred in Quebec City, Canada, which caused 182 declared cases of Legionnaire''s disease and included 13 fatalities. Legionella pneumophila serogroup 1 isolates from 23 patients as well as from 32 cooling towers located in the vicinity of the outbreak were recovered for analysis. In addition, 6 isolates from the 1996 Quebec City outbreak and 4 isolates from patients unrelated to both outbreaks were added to allow comparison. We characterized the isolates using pulsed-field gel electrophoresis, sequence-based typing, and whole genome sequencing. The comparison of patients-isolated strains to cooling tower isolates allowed the identification of the tower that was the source of the outbreak. Legionella pneumophila strain Quebec 2012 was identified as a ST-62 by sequence-based typing methodology. Two new Legionellaceae plasmids were found only in the epidemic strain. The LVH type IV secretion system was found in the 2012 outbreak isolates but not in the ones from the 1996 outbreak and only in half of the contemporary human isolates. The epidemic strains replicated more efficiently and were more cytotoxic to human macrophages than the environmental strains tested. At least four Icm/Dot effectors in the epidemic strains were absent in the environmental strains suggesting that some effectors could impact the intracellular replication in human macrophages. Sequence-based typing and pulsed-field gel electrophoresis combined with whole genome sequencing allowed the identification and the analysis of the causative strain including its likely environmental source.     

Variability of the microfloral makeup of the pathological focus in suppurative infection]

During the period of hospital treatment a change in the composition of would microflora in 145 patients was found to occur, which affected the characteristics of microflora in respect of species, types and strains. This change consisted in the appearance of new (secondary) species, variants and strains and in the disappearance of some original (primary) ones. The populations of staphylococci were found to vary in the level and spectrum of their resistance to antibiotics (resistovars) and in the degree and spectrum of their sensitivity to staphylococcal typing phages (phagovars). Changes in the populations of staphylococci were caused mainly by the cange of S. aureus or by the additional appearance of their new variants which belonged, as a rule, to phage group II, had multiple resistance to antibiotics and corresponded to the phagovars of the hospital flora. The development of hospital variants in open wounds led to an increased general level of resistance of staphylococcal populations to antibiotics.     

Silencing Nrf2 attenuates chronic suppurative otitis media by inhibiting pro-inflammatory cytokine secretion through up-regulating TLR4

Compromised TLR-mediated chronic inflammation contributes to bacterial infection-caused chronic suppurative otitis media, but the mechanisms are unclear. The present study examined the expression status of nuclear erythroid 2-related factor 2 (Nrf2) and TLRs in human middle-ear mucosae tissues collected from patients with chronic suppurative otitis media, chronic otitis media and non-otitis media, and found that Nrf2 was high-expressed, whereas TLR4, instead of other TLRs, was low expressed in chronic suppurative otitis media compared to chronic otitis media and non-chronic otitis media groups. Consistently, inflammatory cytokines were significantly up-regulated in the chronic suppurative otitis media group, instead of the chronic otitis media and non-chronic otitis media groups. Next, LPS-induced acute otitis media and chronic suppurative otitis media models in mice were established, and high levels of inflammatory cytokines were sustained in the mucosae tissues of chronic suppurative otitis media mice compared to the non-otitis media and acute otitis media groups. Interestingly, continuous low-dose LPS stimulation promoted Nrf2 expression, but decreased TLR4 levels in chronic suppurative otitis media mice mucosae. In addition, knock-down of Nrf2 increased TLR4 expression levels in chronic suppurative otitis media mice, and both Nrf2 ablation and TLR4 overexpression inhibited the pro-inflammatory cytokine expression in chronic suppurative otitis media. Finally, we found that both Nrf2 overexpression and TLR4 deficiency promoted chronic inflammation in LPS-induced acute otitis media mice models. Taken together, knock-down of Nrf2 reversed chronic inflammation to attenuate chronic suppurative otitis media by up-regulating TLR4.     

Development of a molecular method for the typing of Brettanomyces bruxellensis (Dekkera bruxellensis) at the strain level

AIMS: In recent years, Brettanomyces/Dekkera bruxellensis has caused increasingly severe quality problems in the wine industry. A typing method at the strain level is needed for a better knowledge of the dispersion and the dynamics of these yeasts from grape to wine. METHODS AND RESULTS: Three molecular tools, namely random-amplified polymorphic DNA, PCR fingerprinting with microsatellite oligonucleotide primers and SAU-PCR, were explored for their relevance to typing strains of Brettanomyces bruxellensis. The results indicated that discrimination of each individual strain was not possible with a single PCR typing technique. We described a typing method for B. bruxellensis based on restriction enzyme analysis and pulse field gel electrophoresis (REA-PFGE). Results showed that electrophoretic profiles were reproducible and specific for each strain under study. CONCLUSIONS: Consequently, REA-PFGE should be considered for the discrimination of B. bruxellensis strains. This technique allowed a fine discrimination of B. bruxellensis, as strains were identified by a particular profile. SIGNIFICANCE AND IMPACT OF THE STUDY: This study constitutes a prerequisite for accurate and appropriate investigations on the diversity of strains throughout the winemaking and ageing process. Such studies will probably give clearer and more up-to-date information on the origin of the presence of Brettanomyces in wine after vinification when they are latent spoilage agents.     

Hearing loss in suppurative meningitis and encephalitis in our cases

Seventeen cases of the suppurative meningitis complicated by hearing loss have been analysed clinically. Emphasis is on the possibility of such a complication in any case of the suppurative infection of CNS. The authors have shown the necessity of otolaryngological examinations in each patient, basing on their own experience and available literature. An early diagnosis and appropriate treatment produces complete recover or decreases the degree of hearing loss being a complication of the suppurative neuroinfections.     

Specificities of bacterial death in culture and suppurative wounds

Electron microscopic autoradiographic study has been performed. In tissue sections, obtained from suppurative wounds, diploforms of cocci and bacilli have been observed. In the former cells DNA and RNA synthesis was normal, however, in the latter cells no synthesis was detected. Such diploforms were not observed in fresh and exhausted cultures. Possible existence in the body of a factor directly injuring bacterial genome is discussed.     

圈养林麝铜绿假单胞菌的分离鉴定及耐药性和病原特性分析

【背景】铜绿假单胞菌感染所致的化脓性疾病是困扰林麝驯养的重要因素,是一类较难防治的细菌性疾病,目前尚无疫苗可用来预防该病。【目的】研究林麝源铜绿假单胞菌的感染现状和分子流行病学规律。【方法】对2014年10月至2015年10月四川宝兴和陕西镇坪两个林麝养殖中心发病林麝中铜绿假单胞菌进行分离鉴定,并对其耐药情况进行分析,利用脉冲场凝胶电泳(PFGE)分型技术研究分离菌的PFGE指纹图谱,探究其流行病学趋势,并对部分菌株的致病性进行分析。【结果】分离得到60株铜绿假单胞菌,其中34株来自镇坪,26株来自宝兴。耐药结果表明:60株林麝源铜绿假单胞菌对17种抗菌药物呈现不同程度耐药性,不同地区和不同样本源间分离的铜绿假单胞菌对17种抗菌药物的耐药性总体趋于一致,多重耐药情况严重,以5耐、6耐为主。分型结果显示:60株铜绿假单胞菌PFGE图谱的相似性为49.1%-100%。经聚类分析得到A-O共15种基因簇,其中优势基因簇为C、E、G、J。致病性结果表明,流行菌株的致病力强弱依次为:动物源菌株环境源菌株,且主要流行菌株(基因簇E、F、J)的致病力大于其余菌群。【结论】铜绿假单胞菌在两地区具有水平传播的途径,本研究可为跨地区林麝化脓性炎症的防控提供理论依据。     

Conventional and molecular approaches to isolates of Salmonella hadar from sporadic and epidemic cases

In September 1994 an outbreak of gastroenteritis occurred in 437 people who had consumed lunch in the canteen of a factory in Central Italy. Salmonella sp. was isolated from stools of 99 patients and in 73 of them Salmonella hadar was identified. This is the first outbreak caused by this serotype described in Italy. In order to examine the genotypic basis of the epidemic strains, molecular typing was applied to sporadic strains isolated before and after the outbreak episode. For this purpose phage type, resistance to antibiotics, DNA plasmid profile and sites of insertion of the mobile element of IS200 were determined. The epidemic strains were genetically distinct from the non-epidemic isolates; they were shown to be phage type 26, harbouring four small plasmids, were resistant to nalidixic acid and showed a unique characteristic IS200 fingerprint. The typing methods used in this study allowed the identification and discrimination of the outbreak strains from related isolates. They can thus be considered as a tool for epidemiological purposes. In addition we should point out the emerging resistance to nalidixic acid, largely used in veterinary medicine, in Salm . hadar .     

Physiological and molecular techniques for the study of bacterial community development in sausage fermentation

The development of the microbial community involved in the production process of Italian dry sausage was investigated using physiological analysis and molecular techniques for strain typing and taxonomical identification. A cycle of sausage production was followed collecting samples during the 2 months of ripening process. Microbiological analysis allowed the identification of the main bacterial groups responsible for the fermentation process as lactobacilli and coagulase-negative staphylococci. The use of a polymerase chain reaction-based technique of strain typing, RAPD fingerprinting, demonstrated that the environmental parameters interact to select a limited number of strains that dominate the fermentation process. The staphylococcal populations were characterized for their physiological properties and the two dominant strains were identified as Staphylococcus xylosus and Staph. sciuri. The use of 16S rDNA sequencing allowed the definition of the taxonomical position of the two dominant strains of lactic acid bacteria, as belonging to Lactobacillus sake and Lact. plantarum.     

化脓性中耳炎病原菌感染的研究进展

化脓性中耳炎是耳鼻喉科临床常见疾病,可导致听力下降、鼓膜充血、鼓膜穿孔、耳鸣、耳痛及流脓等。化脓性中耳炎主要由微生物进入中耳引起感染,使中耳黏膜发生化脓性病变,且不同患者感染的病原菌不同。本文从化脓性中耳炎的发病机制、病原菌及其耐药性和治疗方法等几个方面进行综述,以期为临床化脓性中耳炎的诊断及合理用药提供参考。     

Random amplification of polymorphic DNA (RAPD) of Salmonella: strain differentiation and characterization of amplified sequences

Random amplification of polymorphic DNA (RAPD) was evaluated for its ability to differentiate Salmonella strains from various sources. Under defined conditions RAPD using a 10-mer primer (1254) produced a series of amplification products able to reproducibly distinguish strains representing 20 different serotypes of Salmonella. Primer 1254 also proved capable of discrimination between some but not all isolates of Salm. ser. Enteritidis and Salm. ser. Typhimurium, phage typing proving to be most discriminatory for the latter serotype. Cloning of fragments into a vector allowed sequencing and database searching for identification of fragments and an indication of criteria for primer template interaction in RAPD. Southern blotting using a digoxigenin-labelled probe allowed identification of related bands between RAPD profiles. These observations demonstrate the potential of rapid molecular typing by RAPD for the genomic typing of Salmonella strains.     

Development of a sequence typing scheme for differentiation of Salmonella Enteritidis strains

A DNA sequence typing scheme based on the caiC and SEN0629 loci was developed for differentiation of Salmonella Enteritidis strains and validated using a diverse collection of 102 isolates representing 38 phage types from different sources, year of isolation, geographical locations and epidemiological backgrounds. caiC encodes a probable crotonobetaine/carnitine-CoA ligase, and SEN0629 is a pseudogene. Our system allowed for discrimination of 16 sequence types (STs) among the 102 isolates analysed and intraphage type differentiation. Our findings also suggested that the stability of phage typing may be adversely affected by the occurrence of phage type conversion events. During a confirmatory phage typing analysis performed by a reference laboratory, 13 of 31 S. Enteritidis strains representing nine phage types were assigned phage types that differed from the ones originally determined by the same reference laboratory. It is possible that this phenomenon passes largely unrecognized in reference laboratories performing routine phage typing analyses. Our results demonstrate that phage typing is an unstable system displaying limited reproducibility and that the two-loci sequence typing scheme is highly discriminatory, stable, truly portable and has the potential to become the new gold standard for epidemiological typing of S .?Enteritidis strains.     

Enhanced expression of Duffy antigen in the lungs during suppurative pneumonia.

Duffy antigen is a chemokine binding protein expressed on the surface of erythrocytes and postcapillary venular endothelial cells. It binds selective CXC and CC chemokines with high affinity. Although Duffy antigen is present in the normal pulmonary vascular bed, it is not known whether its expression is altered by innate inflammatory responses in the lungs. We studied Duffy antigen expression by immunohistochemistry in autopsy lung specimens from 16 cases of suppurative pneumonia, 11 cases of acute lung injury, and seven normal lungs. In lungs with suppurative pneumonia, Duffy antigen was expressed in higher numbers of pre- and postcapillary parenchymal vessels compared to normal specimens or specimens with acute lung injury (p<0.03 and p<0.02, respectively). Lungs with suppurative pneumonia also showed Duffy antigen expression on the alveolar septa, whereas this was a rare finding in normal specimens or in acute lung injury (p<0.02). Furthermore, Duffy antigen labeling of the alveolar septa localized to regions with airspace accumulation of neutrophil-rich exudates. In summary, Duffy antigen expression is increased in the vascular beds and alveolar septa of the lung parenchyma during suppurative pneumonia, suggesting that Duffy antigen may have a functional role in the lung parenchyma during inflammation.