Numerical analysis of the characteristics of Corynebacterium diphtheriae strains isolated in Victoria from 1962 to 1971.

A study of the incidence of diphtheria in the State of Victoria, Australia, was carried out. Numerical analysis of the characteristics of 264 strains of Corynebacterium diphtheriae isolated between 1962 and 1971 placed them into 18 varieties plus six strains which were unique in their combination of reactions to the characteristics examined. During the 10-year period, some varieties appeared intermittently and were recognized by certain defining characteristics but exhibited a gradual change in their antigenic structure. In contrast, when the outbreaks were examined over shorter periods of time, a number of varieties and single strains were found which differed greatly from each other yet possessed the same major serotype antigen. These findings are discussed in terms of a ''one-parent'' concept in which the varieties and single strains represent phases of a common ancestor. By inspection and analysis of the characteristics of the strains, certain associations were apparent. For instance, a correlation was found between the antigenic structure of the organism and the colonial appearance on tellurite blood agar. Similarly, correlation was observed between bacteriophage type, diphthericin type and biochemical activity in that a strain which was highly active in one of the properties was also very active in the other two.     

A novel transducible chimeric phage from Escherichia coli O157:H7 Sakai strain encoding Stx1 production

Shiga toxin-producing Escherichia coli (STEC), and especially enterohaemorrhagic E. coli (EHEC) are important, highly virulent zoonotic and food-borne pathogens. The genes encoding their key virulence factors, the Shiga toxins, are distributed by converting bacteriophages, the Stx phages. In this study we isolated a new type of inducible Stx phage carrying the stx1 gene cluster from the prototypic EHEC O157:H7 Sakai strain. The phage showed Podoviridae morphology, and was capable of converting the E. coli K-12 MG1655 strain to Shiga toxin-producing phenotype. The majority of the phage genes originate from the stx2-encoding Sakai prophage Sp5, with major rearrangements in its genome. Beside certain minor recombinations, the genomic region originally containing the stx2 genes in Sp5 was replaced by a region containing six open reading frames from prophage Sp15 including stx1 genes. The rearranged genome, together with the carriage of stx1 genes, the morphology and the capability of lysogenic conversion represent a new type of recombinant Stx1 converting phage from the Sakai strain.     

Towards a subunit vaccine from a Shigella flexneri ΔtolR mutant

Disruption of one or more components of the Tol-Pal system, involved in maintaining the integrity of the outer membrane of Gram-negative bacteria, has been proposed as a method to increase the yield obtained from natural production of outer membrane vesicles (OMV). We present a new OMV-based product, obtained from genetically modified Shigella flexneri 2a with a non-polar deletion in tolR and heat-inactivated (HT-ΔtolR). The S. flexneri ΔtolR strain lead to a higher release of vesicles, more than 8-times when compared to the yield obtained from chemically inactivated wild type strain. S. flexneri mutant strain appeared to be more sensitive to different chemical compounds, including antibiotics, bile salts or human complement and it was also less virulent in both in vitro and in vivo assays. The mutation produced some changes in the LPS O-chain and protein expression. S. flexneri ΔtolR was enriched in long and very long LPS O-chain and expressed a different pattern of surface proteins or lipoproteins. In vitro toxicity and activation properties were determined in Raw 267.4 macrophage cell line. HT-ΔtolR antigenic complex was non-cytotoxic and activation markers, such as MHC-II or CD40, were highly expressed during incubation with this product. Finally, preliminary studies on the antibody response elicited by HT-ΔtolR demonstrated a robust and diverse response in mice. Considering these promising results, HT-ΔtolR antigenic extract appears as a new potential vaccine candidate to face shigellosis.     

Effects of Synbiotic Supplementation on Chronic Inflammation and the Gut Microbiota in Obese Patients with Type 2 Diabetes Mellitus: A Randomized Controlled Study

The aim of this study was to investigate the effects of 24-week synbiotic supplementation on chronic inflammation and the gut microbiota in obese patients with type 2 diabetes. We randomized 88 obese patients with type 2 diabetes to one of two groups for 24 weeks: control or synbiotic (Lacticaseibacillus paracasei strain Shirota (previously Lactobacillus casei strain Shirota) and Bifidobacterium breve strain Yakult, and galactooligosaccharides). The primary endpoint was the change in interleukin-6 from baseline to 24 weeks. Secondary endpoints were evaluation of the gut microbiota in feces and blood, fecal organic acids, high-sensitivity C-reactive protein, lipopolysaccharide-binding protein, and glycemic control. Synbiotic administration for 24 weeks did not significantly affect changes in interleukin-6 from baseline to 24 weeks (0.35 ± 1.99 vs. −0.24 ± 1.75 pg/mL, respectively). Relative to baseline, however, at 24 weeks after synbiotic administration there were positive changes in the counts of Bifidobacterium and total lactobacilli, the relative abundances of Bifidobacterium species such as Bifidobacterium adolescentis and Bifidobacterium pseudocatenulatum, and the concentrations of acetic and butyric acids in feces. No significant changes in inflammatory markers were found in the synbiotic group compared to the control group. However, synbiotic administration at least partially improved the gut environment in obese patients with type 2 diabetes.     

Antibacteriophage action on the larvicidal activity of Bacillus thuringiensis H-14 and Bacillus sphaericus against Culex pipiens

An organically polluted mosquito breeding water was tested for the presence of bacteriophages which could inhibit the larvicidal activity of B. thuringiensis H-14 and B. sphaericus. More than one bacteriophage were isolated which could inhibit the tested bacteria. The sensitivity of the two bacterial species to 12 antibiotics was tested. Two of them, showed no antibacterial action, were selected and considered as antivirus agents in the bacteriophage assays; namely, Amoxycillin and Co-Trimoxazole. Results indicated their antivirus activity as the addition of the antibiotics to the isolated phages could permit normal bacterial growth as well as persistence of larvicidal activity against Culex pipiens larvae.     

Comparative analysis of two bacteriophages of Xanthomonas arboricola pv. juglandis

Walnut blight caused by Xanthomonas arboricola pv. juglandis (Xaj) is one of the most frequent infective diseases of walnut, resulting in serious economic losses. One potential solution to control this disease could be the application of bacteriophages. In this study, 24 phages were isolated from soil and walnut aerial tissues infected with Xaj. Two polyvalent bacteriophages, Xaj2 and Xaj24 were chosen for further characterization including their morphological, physiological and genomic analyses. Xaj2 was classified as Siphoviridae whereas Xaj24 belonged to the Podoviridae family. Both phages demonstrated lytic effect on Xaj in laboratory trials.Complete genomes of Xaj2 and Xaj24 were determined. Genomes of Xaj2 and Xaj24 consisted of 49.241 and 44.861 nucleotides encoding 80 and 53 genes, respectively. Comparative genome analyses have revealed that Xaj2 had a unique genome sequence, while Xaj24 was a phiKMV-like phage and it was most similar to the Prado phage which is virulent for Xylella fastidiosa and Xanthomonas spp.In this study, we present the first two complete Xaj phage sequences enabling an insight into the genomics of Xaj phages.     

Possible effects of antibiotic treatment on the sensitivity and growth requirements of Neisseria gonorrhoeae

The decreased sensitivity to penicillin of Neisseria gonorrhoeae has been a cause of concern during recent years. About 25% of strains sent in for routine diagnosis to the Statens Seruminstitut in 1957 were found to require roughly 20 times as much penicillin to inhibit development completely as was required for strains isolated in 1944. The stability of this reduced sensitivity to penicillin was studied in 20 strains which were subcultured daily on medium without penicillin. A comparison was also made between the sensitivity levels to penicillin and streptomycin for 1957 and 1958. A phenomenon possibly resulting from penicillin therapy of gonorrhoea was noted by the authors in June 1957, when they recorded the appearance of atypical strains having special growth requirements, low viability and weak or absent glucose-fermentation. The incidence, serology and sensitivity to antibiotics of these atypical strains were examined; further research on their different behaviour in response to changes in medium, as compared with that of typical strains, may give rise to some interesting findings.     

Novel Bacteroides host strains for detection of human- and animal-specific bacteriophages in water

Bacteriophages active against specific Bacteroides host strains were shown to be suitable for detection of human faecal pollution. However, the practical application of this finding is limited because some specific host strains were restricted to certain geographic regions. In this study, novel Bacteroides host strains were isolated that discriminate human and animal faecal pollution in Switzerland. Two strains specific for bacteriophages present in human faecal contamination and three strains specific for bacteriophages indicating animal faecal contamination were evaluated. Bacteriophages infecting human strains were exclusively found in human wastewater, whereas animal strains detected bacteriophages only in animal waste. The newly isolated host strains could be used to determine the source of surface and spring water faecal contamination in field situations. Applying the newly isolated host Bacteroides thetaiotaomicron ARABA 84 for detection of bacteriophages allowed the detection of human faecal contamination in spring water.     

Molecular characterization of L class genome segments of a newly isolated turkey arthritis reovirus

Seven strains of turkey arthritis reovirus (TARV) isolated from cases of turkey arthritis were characterized on the basis of their L class genome segment sequences, which were then compared with those of turkey enteric reovirus (TERV) and chicken reovirus (CRV). All three L class gene segments of TARVs and TERVs and their encoded proteins λA, λB, and λC were similar in size to those of CRV reference strain S1133. The conserved motifs such as C2H2 zinc-binding motif and conserved polymerase region were present in λA and λB, respectively. A conserved motif for ATP/GTP-binding site and an S-adenosyl-l-methionine (SAM)-binding pocket for methyltransferase were observed in λC protein of TARVs and TERVs with only one substitution as compared to that in CRV. We propose a new genotype classification system for avian reoviruses (ARVs) based on the nt identity cut-off value for each of the L class. Based on this new genotype classification, all ARVs were divided into six, seven and eight genotypes in L1, L2 and L3 genes, respectively. Interestingly TARVs and TERVs grouped with three CRVs (two arthritic strains from Taiwan and one enteritic strain from Japan) in genotype L1-I and formed a different genotypes (L2-I, L3-I) from CRVs in L2 and L3 genes. The maximum nucleotide divergence was observed in genotypes of L1 and L2 genes but less at amino acid level indicates mostly changes were synonymous type. Compared to L1 and L2 genes, the nonsynonymous changes were more in L3 gene. Point mutations and possible reassortments among TARVs, TERVs and CRVs were also observed.     

Incidence and properties of Staphylococcus aureus associated with turkeys during processing and further-processing operations.

The incidence of Staphylococcus aureus on turkeys sampled at various stages of processing and further-processing was determined on four occasions at each of three different processing plants. For freshly-slaughtered birds, counts from neck skin varied from plant to plant over the range less than 10(2) to greater than 10(5)/g but in all cases the corresponding counts obtained from carcasses sampled after chilling rarely exceeded 10(3)/g and the same was true for samples of mechanically recovered meat (MRM), the final raw product examined. Despite the limited susceptibility of isolates from the different factories to typing by means of either standard human or poultry bacteriophages (55-94% untypable), evidence was obtained with the aid of biotyping for the presence of both human and animal-derived strains. However, some biotypes isolated from MRM were not detected at earlier stages of processing. At one processing plant, an "indigenous'' type of S. aureus was clearly demonstrated. It occurred in high numbers in the defeathering machines (up to 10(5)/swab), was found on carcasses at all subsequent stages of processing over the survey period and was shown to survive routine cleaning and disinfection procedures. Isolates of this type produced unusually large amounts of extracellular "slime'' in artificial culture. Two of the three processing plants yielded isolates which were enterotoxigenic. Of 55 strains from Plant 1, 60% produced enterotoxin C and all were of the "indigenous'' type. In the case of Plant 2, only two type D- and one type F-producing strain were found.     

Toxic Effects Produced by Microcystins from a Natural Cyanobacterial Bloom and a Microcystis aeruginosa Isolated Strain on the Fish Cell Lines RTG-2 and PLHC-1

Toxic cyanobacterial blooms are a worldwide problem, causing serious water pollution and public health hazard to humans and livestock. The intact cells as well as the toxins released after cellular lysis can be responsible for toxic effects in both animals and humans and are actually associated with fish kills. Two fish cell lines—PLHC-1 derived from a hepatocellular carcinoma of the topminnow Poeciliopsis lucida and RTG-2 fibroblast-like cells derived from the gonads of rainbow trout Oncorhynchus mykiss were exposed to several concentrations of extracts from a natural cyanobacterial bloom and a Microcystis aeruginosa–isolated strain. After 24 hours, morphologic and biochemical changes (total protein content, lactate dehydrogenase leakage, neutral red uptake, methathiazole tetrazolium salt metabolization, lysosomal function, and succinate dehydrogenase [SDH] activity) were investigated. The most sensitive end point for both cyanobacterial extracts in PLHC-1 cells was SDH activity, with similar EC₅₀ values (6 μM for the cyanobacterial bloom and 7 μM for the isolated strain). RTG-2 cells were less susceptible according to SDH activity, with their most sensitive end point lysosomal function with an EC₅₀ of 4 μM for the M. aeruginosa–isolated strain and 72 μM for the cyanobacterial bloom. The lysosomal function was stimulated at low concentrations, although SDH activity increased at high doses, indicating lysosomal and energetic alterations. Increased secretion vesicles, rounding effects, decreased cell numbers and size, hydropic degeneration, esteatosis, and apoptosis were observed in the morphologic study. Similar sensitivity to the M. aeruginosa–isolated strain was observed in both cell lines, whereas the cyanobacterial bloom was more toxic to the PLHC-1 cell line.     

Bacteriophage typing of cholera

A bacteriophage-typing scheme for Vibrio cholerae has been developed on the basis of the pattern of susceptibility of V. cholerae strains to four groups of freshly isolated cholera bacteriophages. Some 4066 strains of V. cholerae isolated in Calcutta during the period 1955-61 have been classified into seven types and subtypes. Less than 1% of the strains were untypable. No correlation was found to exist between the phage-types and serological types of V. cholerae. In lysogenic strains, however, a correlation was observed between the phage-types of V. cholerae and their lysogenic phages. Groups of infection deriving from a single source were found to be caused by single phage-types. The author also discusses the practical value of the phage-typing scheme for the epidemiological investigation of cholera.     

Novel prophage-like sequences in Mycoplasma anserisalpingitidis

Mycoplasma anserisalpingitidis is a bacterial waterfowl pathogen. In these days of growing antibiotic resistance, it is necessary to search for alternative methods of defense against Mycoplasma impacts in flocks.In order to identify prophage-like sequences, three established bioinformatics tools (PHASTER, PhiSpy, Prophage Hunter) were used in this study for the in silico screening of 82 M. anserisalpingitidis whole genomes. The VIBRANT software was used as a novel approach to further investigate the possibility of prophages in the sequences.The commonly used softwares found prophage-like sequences in the strains, but the results were inconclusive. The VIBRANT search resulted in multiple hits, and many of them were over 10,000 base pairs (bp). These putative prophages are comparable in size to the few described mycoplasma phages. The translated coding DNA sequences of the putative prophages were checked with protein BLAST. The functions of the proteins found by the BLASTP search are common among bacteriophages. The BLASTN search of the sequences found that many of these were more similar to the M. anatis NCTC 10156 strain, rather than the available M. anserisalpingitidis strains.The initial screening pointed at the presence of novel bacteriophages in the M. anserisalpingitidis and M. anatis strains. The VIBRANT search results were very similar to each other and none of these sequences were part of the core genome of M. anserisalpingitidis, with a few exceptions. The VIBRANT analysis explored presumably intact, novel prophages.     

Epidemiology of Staphylococcus aureus in patients with cystic fibrosis

Seven hundred and thirty-four isolates of Staphylococcus aureus, recovered from the sputum of 238 cystic fibrosis patients in six French hospitals, were characterized by esterase electrophoretic typing, capsular polysaccharide sero-typing and phage typing and tested against 14 antibiotics for sensitivity. Thirty-four esterase electrophoretic types were found with a genotypic diversity coefficient of 0·91. Five hundred and forty-eight (78·7%) isolates produced capsular polysaccharide and 350 (50·3%) were type 8. Four hundred and sixty isolates (66·6%) were phage typable and 202 (28·2%) were lysed by group III bacteriophages. No esterase electrophoretic type, capsular type or phage type was specific to cystic fibrosis. Isolates belonged to a wide range of types, similar to strains acquired outside hospitals. Eighty-five patients had three or more consecutive isolates over at least 6 months. The ability of S. aureus to persist for long periods of time has been demonstarated in 73% of them. Methicillin-resistance was enacountered among 73 strains (9·8%) which were also multiresistant. Two hundred and eighty-nine (39·9%) strains were sensitive to all antibiotics tested except to penicillin. Pristinamycin and co-trimoxazole were the most effective antibiotics. These results could contribute to the elaboration of a rational approach to the prophylaxis and therapy of respiratory staphylococcal infections in cystic fibrosis patients.     

珠海登革病毒分离株的型别鉴定与序列分析研究

[目的]了解珠海市部分人群和入境船员中登革热的流行情况和珠海市流行的登革病毒株的型别和序列特征。[方法]分别采用胶体金快速法、酶联免疫吸附试验(ELISA)对登革热抗体进行检测；采用细胞培养和逆转录一聚合酶链反应(RT-PCR)技术进行病毒鉴定,并对其结构蛋白基因序列进行克隆、测序及同源性分析。[结果]来自东南亚国家船舶的船员血清中登革病毒IgM抗体检出率为4．40％,IgG抗体为20．33％,其阳性样本经RT-PCR扩增,结果为阴性。而对本室保存的2001年珠海散发的登革热患者阳性血清,经用C6/36细胞培养分离出1株登革病毒,用登革病毒通用引物和Ⅱ型特异性引物,扩增出相应的片段,证实为登革Ⅱ型病毒；该分离株(DV2-1)的基因序列与其他9株登革Ⅱ型病毒进行系统发育关系的分析,结果显示分离株(DV2-1)与GD08/98株、FJ11/99株、16681株位于相近的分支,其中与1998年广东流行株GD08/98株系统进化关系最近。[结论]2001年珠海地区登革热的散发流行为登革Ⅱ型病毒所致,推测其可能是输入性传染。     

Egg-related Salmonella enteritidis, Italy, 1991

In recent years, Salmonella enteritidis has become an increasingly important public health problem in Italy. In some parts of the country, the fraction of total human salmonella isolates accounted for by S. enteritidis has risen from 3-4% in the mid-1980s to more than 30% in 1990. Between 1990 and 1991, the number of reported S. enteritidis outbreaks increased more than sixfold. The 33 outbreaks reported in 1991 occurred in seven contiguous regions in northern and central Italy and were clustered in time between June and October; in the majority, products containing raw or undercooked shell eggs were implicated. Five of the egg-related outbreaks that occurred within a 30 kilometre radius over a 7-week period were investigated in detail. A phage type 1 strain containing a 38·9 MDa plasmid appeared responsible for three of the outbreaks, while in the remaining two a phage type 4 strain, also with a 38·9 MDa plasmid was isolated. Efforts are being made to enhance epidemiological surveillance and laboratory evaluation, and the use of pasteurized eggs has been recommended for high-risk populations.     

Intermittent administration of a fasting-mimicking diet intervenes in diabetes progression,restores β cells and reconstructs gut microbiota in mice

Fasting and especially intermittent fasting have been shown to be an effective intervention in many diseases, such as obesity and diabetes. The fasting-mimicking diet (FMD) has recently been found to ameliorate metabolic disorders. To investigate the effect of a new type of low-protein low-carbohydrate FMD on diabetes, we tested an FMD in db/db mice, a genetic model of type 2 diabetes. The diet was administered every other week for a total of 8 weeks. The intermittent FMD normalized blood glucose levels in db/db mice, with significant improvements in insulin sensitivity and β cell function. The FMD also reduced hepatic steatosis in the mice. Deterioration of pancreatic islets and the loss of β cells in the diabetic mice were prevented by the FMD. The expression of β cell progenitor marker Ngn3 was increased by the FMD. In addition, the FMD led to the reconstruction of gut microbiota. Intermittent application of the FMD increased the genera of Parabacteroides and Blautia while reducing Prevotellaceae, Alistipes and Ruminococcaceae. The changes in these bacteria were also correlated with the fasting blood glucose levels of the mice. Furthermore, intermittent FMD was able to reduce fasting blood glucose level and increase β cells in STZ-induced type 1 diabetic mouse model. In conclusion, our study provides evidence that the intermittent application of an FMD is able to effectively intervene in the progression of diabetes in mice.     

Properties of an enzyme-based low-level iodine disinfectant

Strains of Pseudomonas stutzeri developed stable resistance to chlorhexidine diacetate (CHA) or cetylpyridinium chloride (CPC) when exposed to gradually increasing concentrations of either antibacterial agent. Such strains showed reduced sensitivity to other non-antibiotics, including triclosan, and to some antibiotics, although this varied from strain to strain. Resistant strains were inactivated less readily by CHA or CPC and were less sensitive to sodium dodecyl sulphate. Some CHA-resistant and some CPC-resistant strains were more hydrophobic than the parent strains. Alterations in the cell envelope are likely to be responsible for non-specific changes in sensitivity to several antibacterial agents. Attempts to transfer CHA or CPC resistance by conjugation were unsuccessful. DNA from some CHA- or CPC-resistant strains could transform Ps. stutzeri strain JM 302, a histidine auxotroph, to prototrophy.     

An experimental study on the usefulness of bacteriophage in the prophylaxis and treatment of cholera

Five different samples of bacteriophages were tried on weanling rabbits intra-intestinally infected with the Inaba 569 B strain of Vibrio cholerae. Three of these prevented the onset of the disease and proved useful in the treatment of early cases of experimental cholera. The prophylactic and curative (if administered early) values of choleraphage observed during these studies appear to agree with the observations of several workers in clinical cases. One of the choleraphages used proved to be valuable both prophylactically and therapeutically in experimental cholera as well as in clinical cases. It was concluded that bacteriophage was most useful in the prevention of the disease, but that therapeutically its beneficial effect could be obtained only if treatment were initiated at the early stage.     

Biotype traits and antibiotic susceptibility of Vibrio cholerae serogroup O1 before, during and after the emergence of the O139 serogroup.

Sixty-nine strains of Vibrio cholerae O1 isolated at different times were analysed to investigate if there were any differences among the O1 strains isolated before, during and after the advent of the O139 serogroup. Of the 69 O1 strains examined, 68 belonged to the Ogawa serotype while one belonged to the Inaba serotype. With the exception of one strain all other strains of V. cholerae O1 belonged to the eltor biotype. A single O1 strain isolated before the emergence of the O139 serogroup could not be classified as either eltor or classical biotype because it was resistant to both classical and eltor specific bacteriophages. Marked variations in the susceptibility to antibiotics of V. cholerae O1 isolated during the different periods were observed. In addition, strains of V. cholerae isolated after the epidemic of serogroup O139 in Calcutta showed an expanding R-type with resistance to a variety of drugs as compared to the O1 strains isolated before the advent of the O139 serogroup. From this study, it is clear that there is a substantial mobility in genetic elements of V. cholerae O1 which necessitates a continuous monitoring to keep abreast of the changing traits of the etiologic agent of cholera.