Materials forPythium flora of Japan. X. Occurrence,identification and seasonality ofPythium spp. in three pond waters and mud soils in Osaka

Seasonal fluctuations ofPythium species in three pond waters and mud soils in Sakai, Osaka, were studied for periods of two years and 14 months, respectively. Thirteen species and four groups were isolated from the pond waters while nine species and three groups were obtained from the mud soils. The highest frequency and total number of species were recorded in the cold and moderately warm seasons and the lowest in summer. Most of the species isolated from the pond waters have been previously recorded from aquatic habitats.     

Effect of temperature,hydrogen ion concentration and osmotic potential on oospore germination of fivePythium spp. isolated from pond water

Oospore germination occurred over a temperature ranging of 15–35°C forPythium coloratum, 10–35°C forP. diclinum, 15–30°C forP. dissotocum, 7–30°C forP. monospermum, and 10–30°C forP. pleroticum. Optimum temperature was 25°C for all species tested. In case of pH, oospore germination occurred over a range of 4.76–8.55 with an optimum of 6.40–7.40. The least germination occurred at pH 4.76 forP. coloratum, P. diclinum, P. monospermum andP. pleroticum, whileP. dissotocum germinated from pH 5.02. Oospores of the all tested pythia were able to germinate at –0.13 to –1.65 MPa and could not germinate at –3.40 MPa, with the highest germination rate at –0.27 to –0.47 MPa. The effect of temperature, pH and osmotic potential on oospore germination was discussed in relation to pollution of pond water.     

A technique for isolating bacteria-freePythium spp. from pond water

A special technique for isolating aquaticPythium spp. that are free from bacteria is presented and discussed by citing the disadvantages of the previous studies.     

Effect of temperature,hydrogen ion concentration and osmotic potential on zoospore production by threePythium species isolated from pond water

Pythium fluminum produced zoospores most abundantly at 15°C, whereas the optima forPythium group F andP. marsipium were 20 and 25°C, respectively. Increasing the incubation temperature above the optimum resulted in the decrease of the duration of zoospore production. InPythium group F the ability to produce zoospores was not lost even after incubation at 40°C for 24 h. On the other hand,P. marsipium andP. fluminum lost the ability under these conditions. Zoospore production was inhibited at pH 4.5 and 10.5 in all the species tested.Pythium fluminum andP. marsipium were found to have two pH optima for zoospore production (7.5 and 9.5 for the former and 5.5 and 8.5 for the latter). The optimum pH for zoospore production byPythium group F was 6.5–7.5. Moderate osmotic potentials (–0.27–0.47 MPa) appeared to favor zoospore production by the pythia tested. The effect of temperature, pH and osmotic potential on zoospore production was discussed in relation to pollution of pond water.     

Virulence and antibiotic susceptibility of Aeromonas spp. isolated from drinking water

Aeromonas isolates from tap water, mineral water, and artesian well water were investigated for their ability to produce different potential virulence factors or markers such as hemolysins, cytotoxins, phospholipase, DNase, hydrophobicity and their ability to adhere to epithelial cells and to abiotic surfaces. The susceptibility to antibiotics of Aeromonas isolates was also examined. Majority of the isolates displayed hemolytic activity against sheep erythrocytes, while only 7 of the 23 Aeromonas strains displayed DNase activity and 4 of the 23 Aeromonas strains tested were regarded as positive for phospholipase production. Most of the isolates showed cytotoxic activities in culture filtrate dilutions at titer of 1/8 or lower. No general relation between the strain isolated and the ability to interact with epithelial cells could be established. Using the bacterial adherence to hydrocarbons method, most of the strains were classified as highly hydrophilic. All five Aeromonas jandaei strains isolates, 9 of the 12 Aeromonas sp strains and four of the five Aeromonas hydrophila were multidrug resistant. The most active antimicrobial was ciprofloxacin (susceptible in 100% of the isolates), and the least active antibiotic was ampicillin (resistance in 92% of the isolates). The majority of the isolates tested were not killed by chlorine at 1.2 mg/l. Whether the high tolerance to chlorine of Aeromonas isolates can be linked to greater virulence is not know.     

Comparison of different media for the identification and quantification ofAeromonas spp. in water

In order to assess the suitability of the Starch glutamate ampicillin penicillin-10C agar for the isolation ofAeromonas spp. from waters it was necessary to compare the properties of this medium with those of three others, Starch ampicillin agar, Ampicillin dextrin agar and m-Aeromonas medium, and to monitor different kinds of waters. A selection of forty eight samples were taken from moderately polluted river water, highly polluted river water, polluted sea water (littoral) and treatment & distribution water and monitored using these media. The results were similar with Ampicillin dextrin agar, m-Aeromonas medium and Starch glutamate ampicillin penicillin-10C, but the simplicity of composition and use and its selectivity recommends the last medium as the most adequate for the isolation ofAeromonas spp.Abbreviations ADA ampicillin dextrin agar - mA m-aeromonas medium - SA starch ampicillin agar - SGAP-10C starch glutamate ampicillin penicillin-10C     

Virulence and antimicrobial resistance potential of Aeromonas spp. associated with shellfish

Aeromonas spp. are associated with seafood-related outbreaks worldwide. In seafood industry, shellfish play a major role in global seafood production. With this emerging trend of shellfish consumption, shellfish-related bacterial infections are being reported frequently. Aeromonas spp. are natural contaminants found in shellfish. Although 36 species have been identified, some species including Aeromonas hydrophila, Aeromonas caviae and Aeromonas veronii biotype sobria have dragged major attention as foodborne pathogenic bacteria. The ability to elaborate a variety of virulence factors of Aeromonas spp. contributes to the pathogenic activities. Also, emerging antimicrobial resistance in Aeromonas spp. has become a huge challenge in seafood industry. Furthermore, multidrug resistance increases the risk of consumer health. Studies have supplied pieces of evidence about the emerging health risk of Aeromonas spp. isolated from seafood. Therefore, the present review was intended to highlight the prevalence, virulence and antimicrobial resistance of Aeromonas spp. isolated from various types of shellfish.     

志贺菌毒力大质粒大片段敲除方法的建立

目的:构建志贺菌毒力大质粒大片段缺失突变体库。方法:首先利用λ-Red重组系统构建弗氏2a志贺菌301株毒力大质粒特定位点缺失株,再在距离此位点20 kb处缺失另一突变位点,最后根据重组酶识别远端FRT位点的特性,将两个远端FRT位点之间的DNA序列全部缺失。结果:敲除了毒力大质粒24 kb的DNA序列。结论:利用λ-Red重组系统及FLP-FRT位点特异性识别重组系统可以对志贺菌毒力大质粒逐步进行大片段的敲除,构建大质粒大片段缺失突变体库。     

Occurrence ofDinophysis spp. and toxic shellfish in the Northern Adriatic

The dynamics of the toxicity of the musselMytilus galloprovincialis was compared between two different shellfish farms, 5 km apart, but using the same cultivation technique. The main differences concerned the freshwater influx and the open aspect to the Gulf of Trieste. It is suggested that a deep closed bay and abundant fresh water inflow are the two main conditions for the low toxicity levels in mussels and for shorter periods of danger. A detailed study of the phytoplankton samples revealed the presence of eight species ofDinophysis in the area of both shellfish farms. During the period of the DSP outbreak in Slovenia (autumn and winter 1989).D. fortii andD. acuminata were the most frequentDinophysis species. There was a high positive correlation between the onset of mussel toxicity and the appearance ofDinophysis spp.     

In Vitro Evaluation of Putative Virulence Attributes of Oral Isolates of <Emphasis Type="Italic">Candida</Emphasis> spp. Obtained from Elderly Healthy Individuals

Identification of Candida isolates obtained from oral cavity of elderly healthy individuals revealed the predominance of non-albicans Candida species (88.9%) compared to Candida albicans (11%). CHROMagar Candida differential medium and PCR revealed the presence of Candida tropicalis (33.3%), Candida glabrata (27.8%), and Candida krusei (16.7%). We investigated the presence of virulence attributes in a total of 18 isolates, including acid protease and phospholipase production, hemolytic activity, and biofilm production. Extracellular protease was found in five isolates (27.8%) whereas extracellular phospholipase was found in three isolates (17%). All isolates showed hemolytic activity. About 56% of the isolates were weakly positive for biofilm formation (score +) whereas a minority (5.6%) of them showed strong biofilm formation (score 4+). Susceptibility in vitro of the isolates to fluconazole was carried out by microdilution method. Fluconazole showed a strong inhibition against most buccal isolates. The resistant isolates were 2 C. tropicalis, 2 C. glabrata, and 1 C. krusei.     

High frequency of hemolytic and cytotoxic activity in Aeromonas spp. isolated from clinical, food and environmental in Rio de Janeiro, Brazil

Molecular study of aerolysin and cytotonic enterotoxin genes by PCR and colony blot hybridization was performed in 117 strains of Aeromonas spp. isolated from different sources. Homogeneous distribution of these genes in A. hydrophila complex strains was observed. For A. caviae and A. sobria complex strains, aerolysin genes were more frequent than cytotonic enterotoxins genes. Of 64 A. caviae complex strains, only one (1.5%) amplified the 451 bp product for the aer gene, however, the same primers detected a 400 bp product in 50 (78%) strains. This product was sequenced and had two short regions with homology to several hemolysin genes. The genotype aer ⁺/aerA⁺/hly ⁺/ast ⁺/alt ⁺ was detected in six A. hydrophila strains from food and environmental source. The most common genotype found in A. hydrophila strains was hly ⁺ (85%) and aerA⁺ (78.7%), while in A. caviae complex strains was aerA⁺ (32.8%). All A. veronii complex sobria strains were aer ⁺/aerA⁺. All A. caviae and A. hydrophila were positive when tested with aer probe using the colony blot test. Thirty-seven percent of A. hydrophila and 53% of A. caviae tested were positive for ast probe. Eighty-nine percent of samples were cytotoxic in Vero cells. Our data demonstrated that Aeromonas spp. can harbor and express virulence genes and reinforce the potential of Aeromonas as a human pathogen.     

Using the Tools at Hand: Manual Laterality and Elementary Technology in Cebus spp. and Pan spp.

Cebus and Pan appear to be a remarkable example of evolutionary convergence in behavioral ecology. We examine their apparently analogous solutions to problems posed by laterality of hand function and elementary technology. We scrutinize appropriate published data in a meta-analysis, focusing on Cebus apella and C. capucinus and on Pan paniscus and P. troglodytes. We compare behavioral data in terms of captive versus wild, and tool use versus non-tool use, but notable gaps exist in the data, especially for bonobos. Cebus and Pan spp. are equivalent tool users in captivity, but chimpanzees are notably more extensively so in nature. For hand preference, captive bonobos and wild and captive chimpanzees show ambipreference for non-tool-use patterns. For both Cebus spp. and Pan spp., there is a tendency for individuals to be committed exclusively to one hand or the other for tool use. The data for laterality of hand function fit consistently into the five-level model proposed by McGrew and Marchant (1996).     

Increased resistance to antifungal antibiotics ofCandida spp. adhered to silicone

Summary The minimal inhibitory concentrations (MICs) and minimal fungicidal concentrations (MFCs) of amphotericin B, miconazole, ketoconazole, fluconazole, and itraconazole were determined for non-adhered cells and cells adhered to sections of a silicone urinary catheter. The densities of adhered cells were established with cells radiolabeled with tritiated leucine. Well defined MICs and MFCs were established for amphotericin B for representative adhered strains. In contrast, the azoles, especially fluconazole, did not give clear end points and the MICs and MFCs were arbitrarily determined. MFCs for the adhered cells generally were 2- to 5-fold higher than those of non-adhered cells. Techniques that include adhered-cell susceptibilities may be necessary before antifungal regimens for prosthetic device-associated yeast infections are appropriately defined.This paper is dedicated to Professor Herman Jan Phaff in honor of his 50 years of active research which still continues.     

Extracellular enzymatic activity of Malassezia spp. isolates

Extracellular enzymatic activity of different species of Malassezia spp was evaluated. Thirty-three isolates of animal origin (dogs and cats) and stock culture samples were studied. Twenty isolates of M. pachydermatis, 8 of M. furfur, 2 of M. sympodialis and M. globosa and one of M. restricta, M. obtusa and M. slooffiae were examined. The enzymatic activity was investigatedusing Api Zym system. The enzymatic patterns showed light differences. Esterase lipase, Phosphatase acid and Naphtol-AS-BI-phosphohydrolase were produced in significant amounts from most isolates excepted for M. restricta, confirming the limited enzymatic activity of this species. Data obtained from the other new species described after the revision of the genus, appear to be quite homogeneous. Dixon’s broth appeared to be a valid medium for the growth of all Malassezia spp. This revised version was published online in June 2006 with corrections to the Cover Date.     

Antagonism of Bacillus spp. Isolated from Marine Biofilms Against Terrestrial Phytopathogenic Fungi

We aimed at determining the antagonistic behavior of bacteria derived from marine biofilms against terrestrial phytopathogenic fungi. Some bacteria closely related to Bacillus mojavensis (three isolates) and Bacillus firmus (one isolate) displayed antagonistic activity against Colletotrichum gloeosporioides ATCC 42374, selected as first screen organism. The four isolates were further quantitatively tested against C. gloeosporioides, Colletotrichum fragariae, and Fusarium oxysporum on two culture media, potato dextrose agar (PDA) and a marine medium-based agar [yeast extract agar (YEA)] at different times of growth of the antagonists (early, co-inoculation with the pathogen and late). Overall antagonistic assays showed differential susceptibility among the pathogens as a function of the type of culture media and time of colonization (P < 0.05). In general, higher suppressive activities were recorded for assays performed on YEA than on PDA; and also when the antagonists were allowed to grow 24 h earlier than the pathogen. F. oxysporum was the most resistant fungus while the most sensitive was C. gloeosporioides ATCC 42374. Significant differences in antagonistic activity (P < 0.05) were found between the different isolates. In general, Bacillus sp. MC3B-22 displayed a greater antagonistic effect than the commercial biocontrol strain Bacillus subtilis G03 (Kodiak). Further incubation studies and scanning electronic microscopy revealed that Bacillus sp. MC3B-22 was able to colonize, multiply, and inhibit C. gloeosporioides ATCC 42374 when tested in a mango leaf assay, showing its potential for fungal biocontrol. Additional studies are required to definitively identify the active isolates and to determine their mode of antifungal action, safety, and biocompatibility.     

节杆菌环境适应性的基因组学研究进展

节杆菌分布广泛,能适应多种环境条件,而且多数节杆菌具有营养多功能性,能降解多种环境污染物,因而受到人们的广泛关注。近年来,随着多株节杆菌基因组的测序完成,人们对节杆菌环境适应性的分子机制有了全面的认识。基因组学研究结果表明,节杆菌在σ因子、氧化应激、渗透应激、饥饿应激、温度应激等胁迫应激反应相关基因方面的特点使其能够在多种环境条件下生存。本文挑选部分具有代表性的节杆菌基因组学研究,对其环境适应性的基因组学基础进行综述,以期为利用节杆菌进行环境污染修复提供理论基础,并为其它细菌的环境适应性机制研究提供参考。     

Colony interactions and mycoparasitism betweenAcremonium furcatum andAspergillus spp.

Summary Colony interactions and mycoparasitism betweenAcremonium furcatum and ten species of Aspergillus were studied in dual cultures. It was observed thatA. furcatum overgrew the colonies of all the ten species of Aspergillus studied. During the mycoparasitic interactions frequent coiling, penetration and growth of hyphae ofA. furcatum inside the conidiophores of six species of Aspergillus were observed.     

Analysis of extracellular proteins of two Perkinsus spp. isolated from the softshell clam Mya arenaria in vitro

Biochemical characterization of the extracellular proteins (ECP) of two softshell clam Perkinsus spp. cloned isolates, Perkinsus chesapeaki isolate G-117 and Perkinsus marinus H-49, was performed and compared to that of the oyster-derived P. marinus isolate P-1. G-117 and H-49 demonstrated distinct differences in enzyme activities; however, all three isolates shared common bands. Substrate-impregnated gels showed H-49 to possess proteolytic activities while G-117 did not. Inhibition studies revealed that H-49 ECP contain serine proteases similar to those described for P-1. The G-117 ECP lacked proteolytic activity but showed a higher production of lipolytic enzymes than H-49 or P-1. Optimal in vitro growth temperatures for the two clam isolates were generally lower than those for P-1. G-117 showed faster growth at lower salinities than either H-49 or P-1. Clam Perkinsus spp. isolates appear to be better adapted to lower salinities and temperatures than the P. marinus isolate of the eastern oyster.     

蝴蝶兰几丁质酶基因的克隆及表达特性分析

利用RT-PCR及RACE技术,克隆到蝴蝶兰1个几丁质酶基因PhCHT(GenBank登录号为KT992851),该基因cDNA全长1 210bp,包含37bp的5′-UTR、933bp开放阅读框和240bp 3′-UTR,编码310个氨基酸;该蛋白为糖苷水解酶第19家族成员,兼具有溶菌酶活性;生物信息学分析显示,该蛋白具N-端信号肽和跨膜结构,为胞外分泌蛋白;该蛋白与海枣、谷子、油棕和拟南芥的几丁质酶类似蛋白相近,并且在系统进化树上与甘蔗和陆地棉的Ⅶ类几丁质酶同属一个分支。PhCHT基因的表达分析表明,PhCHT在蝴蝶兰营养器官和生殖器官中均有表达,根中表达量最高;13℃/8℃低温处理3、6、9和15d时该基因的表达被抑制,4℃低温处理1、2和4h表达量升高。研究表明,PhCHT基因能够响应短期的冷胁迫。研究结果为进一步研究蝴蝶兰几丁质酶的系统进化及抗性育种奠定了基础。     

Studies on five species of Pythium parasitic on mustard and cabbage in India

Diseased seedlings and roots of cabbage and mustard of Khurpatal (Nainital), were surveyed for zoosporic plant pathogens. Pythium aphanidermatum and P. debaryanum were isolated from cabbage and P. middletonii, P. spinosum and P. undulatum were isolated from mustard. Pathogenicity tests conducted in the glasshouse indicated that P. debaryanum and P. spinosum are the most virulent pathogens of cabbage and mustard respectively.