SEM morphological survey on the egg shell in the italian Anostracans (Crustacea,Branchiopoda)

The screening of the egg shell morphology of all italian species of Anostracans (14) by means of SEM, as well as the analysis of preliminary data on electrophoretical patterns of the adults, concerning the genus Chirocephalus (five species), suggest the need for a reconsideration, from a taxonomical point of view, especially as far as the species of the diaphanus group (Ch. diaphanus, Ch. ruffoi, Ch. salinus, Ch. sibyllae) are concerned.In this light, within the above mentioned group remarkable differences have been observed for Ch. ruffoi, whilst for other species like Ch. diaphanus and Ch. sibyllae, conspecificity is suspected. Electrophoretical and reproductive isolation studies, in progress, will provide a clearer understanding of the actual situation.Research carried out with M.P.I. (60%) and C.N.R. (chBr. 83.02549.04 Gruppo di Biologia Naturalistica) financial support.Research carried out with M.P.I. (60%) and C.N.R. (chBr. 83.02549.04 Gruppo di Biologia Naturalistica) financial support.     

Characterization of Natronobacterium magadii phage ΦCh1, a unique archaeal phage containing DNA and RNA

A novel archaeal bacteriophage, ΦCh1, was isolated from a haloalkalophilic archaeon Natronobacterium magadii upon spontaneous lysis. The phage-cured strain N. magadii (L13) was used to demonstrate infectivity of phage ΦCh1. The turbid-plaque morphology and the fact that N. magadii cells isolated from plaques were able to produce phage indicated that ΦCh1 is a temperate phage. The phage morphology resembles other members of Myoviridae -infecting Halobacterium species. In solution below 2 M NaCl, the phage lost its morphological stability and infectivity. One- and two-dimensional SDS–PAGE of phage particles revealed at least four major and five minor proteins with molecular masses ranging from 15 to 80 kDa and acidic isoelectric points. Southern blot analysis of chromosomal DNA of a lysogenic N. magadii strain showed that ΦCh1 exists as a chromosomally integrated prophage. The phage particles contain both double-stranded, linear DNA (approx. 55 kbp) as well as several RNA species (80–700 nucleotides). Hybridization of labelled RNA fragments to total DNA from N. magadii and ΦCh1 showed that the virion-associated RNA is host encoded. Part of the phage DNA population is modified and restriction analysis revealed evidence for adenine methylation. Phage ΦCh1 is the first virus described for the genus Natronobacterium , and the first phage containing DNA and RNA in mature phage particles.     

Keratinophilic fungi on the hair of cows,donkeys, rabbits,cats, and dogs from the West Bank of Jordan

The mycoflora of hair of 111 cows, donkeys, rabbits, cats, and dogs were analysed and the frequency of occurrence and the relative importance value of the different fungal species isolated were calculated. Total numbers of species 56, 45, 48, 23, and 11 were recovered from cows, donkeys, rabbits, cats, and dogs respectively. The majority of the keratinophilic fungi isolated were either well-known mycotic agents or have been recovered from various animal and human lesions. These comprised 87.8%, 66.7%, 61.4%, 59.3%, and 55.2% of the hair mycoflora in cats, dogs, rabbis, cows, and donkeys respectively. Seven species of dermatophytes were isolated: Trichophyton mentagrophytes, T. verrucosum, Microsporum gypseum, M. nanum, T. ajelloi, and M. canis. Cats harboured the largest number of dermatophytes (5 species), followed by cows (4 spp), rabbits (3 spp), and donkeys and dogs (1 sp). The role of these animals in the persistence and transmission of pathogenic fungi is discussed.     

Structural characterization of adrenal chromogranin A and parathyroid secretory protein-I as homologs

We have isolated and purified adrenal chromogranin A (Ch A) for the purpose of making structural comparisons to parathyroid secretory protein-I (SP-I), because our earlier data indicated these two molecules may be the same protein. An improved purification step, using high-performance liquid chromatography (HPLC), has enabled us to demonstrate that both SP-I and Ch A consists of two species, one of approximately 72,000 Da and one of approximately 66,000 Da. The amino acid composition is the same for all four species. The difference in molecular mass is assumed to be due to carbohydrate content. Cyanogen bromide digestion of each of the four samples, followed by HPLC separation of the generated peptides, resulted in a chromatographic profile that was the same for each digest. Amino acid analysis of the eight peptide fragments obtained from each digest indicates that both species of Ch A and both species of SP-I yielded the same peptide mixtures following this cleavage reaction. One large (approximately 50,000 Da) CNBr peptide was obtained and seven smaller ones, one of which contains cysteine. The large fragment behaved similarly to the intact molecule in a radioimmunoassay. HPLC separation of tryptic digests of Ch A (72,000 Da) and SP-I (72,000 Da) also resulted in elution profiles that were very similar to each other. Amino acid analysis revealed 23 peptides common to each digest. Ch A contained four peptides ranging in size from 4 to 30 residues that were not observed in the SP-I digest. SP-I contained two peptides, each with about 30 residues, that were not found in the Ch A digest. Nothing unusual was noted in any of the uncommon peptides. Thus, both a chemical and an enzymatic digestion of these molecules followed by analysis of the peptides generated, indicates that SP-I and Ch A are nearly identical homologs.     

Keratinophilic fungi isolated from hospital dust and soils of public places at Gulbarga,India

Keratinophilic fungi were isolated from corridor dust of 11 hospitals and soils of 21 public places using a hair baiting technique. A total of 41 species belonging to 24 genera were recovered. Among the dermatophytes and related species, Microsporum gypseum was predominant, followed by Chrysosporium keratinophilum, Trichophyton mentagrophytes, M. nanum and Ch. tropicum. Other species were represented by 32 species belonging to 21 genera. Most of the species isolated are known to be agents of human and animal infection or have been isolated from human and animal lesions. The fungi observed here are discussed in relation to their global distribution.     

Environmental mycobacteria in Korea. I. Distribution of the organisms

Environmental mycobacteria in Korea have been investigated by examining 54 soil, 111 house dust, 63 well water, and 98 sewage samples collected from 123 randomly selected areas in Korea during the fourth nationwide tuberculosis prevalence survey in 1980. A variety of mycobacteria were isolated from 76% of soil, 67% of sewage, 43% of well water, and 7% of house dust samples. Some samples yielded more than one species; thus 56 strains were obtained from soil, 107 strains from sewage, 48 strains from well water and 8 strains from house dust. Mycobacterium fortuitum was the most common species of environmental mycobacteria in Korea and the species was distributed equally in all types of samples tested. The M. terrae complex was also one of the common species of environmental mycobacteria and it seemed to be more abundant in water samples than in soil. Scotochromogenic slow growers M. scrofulaceum and M. gordonae were common microbes in soil and water samples, although the latter was more frequently detected in water samples. Scotochromogenic rapid growers M. flavescens and M.phlei, and photochromogenic rapid grower M. vaccae were isolated more frequently from sewage or water samples than from soil. Nonphotochromogenic rapid growers M. chelonei (chelonae) and M. smegmatis were isolated mostly from sewage and the former was rarely found in soil and well water samples. The clinically important species M. avium-intracellulare complex was found less frequently in all types of test samples.     

中国蜡梅属植物过氧化物同工酶的研究

采用聚丙烯酰胺凝胶电泳系统,进行了蜡梅属8种植物过氧化物同工酶的研究。结果发现：该属8种植物的酶谱差异显著,每个种均有其特征酶谱。根据其种的酶谱酶带数目、Rf、宽度及其活性强弱的不同,可将各个种加以区别,但不支持保康蜡梅和突托蜡梅作为独立种的存在,也不支持蜡梅属分为蜡梅组和新蜡梅组的观点。同时,采用排序方法对蜡梅属8种植物酶谱的相似程度进行排序结果发现：与该属8种植物酶谱分类相吻合,并从分子水平上论证了它们的亲缘关系,即保康蜡梅为蜡梅的种内变异,以作蜡梅栽培品种处理;突托蜡梅作为浙江蜡梅的变种为佳,从而为蜡梅属植物种群的合理划分,提出了新的依据和手段。     

Isolation of keratinophilic fungi from floors in Roman primary schools

Keratinophilic fungi are present in the environment with variable distribution patterns that depend on different factors, one of which, of fundamental importance, is human and or animal presence. The present study was conducted in the environment and classrooms of schools in order to evaluate the relationship between the human presence and the presence of keratinophilic fungi. In order to achieve this goal, a new isolation technique was used. From 20 samples, 253 colonies of keratinophilic fungi were isolated. The results showed that species of the genus Chrysosporium were present in 100% of the samples, while Microsporum and Trichophyton species were present in 40% and 65% of the samples respectively. The percentage of three pathogenic species, M. canis (25), T. mentagrophytes (10) and M. gypseum (10) was significant. The other species isolated were: T. terrestre (55%), Trichophyton sp. (35%), M. cookei (25%) and T. ajelloi (10%). A correlation between the amount of gathered dust and the number of colonies of keratinophilic fungi isolated was not found.     

Dermatophytes isolated from patients in University Hospital,Kuala Lumpur,Malaysia

A total of 576 dermatophytes were isolated from patients with a variety of skin infections from January 1993 to May 2000. Ten species of dermatophytes were identified: Epidermophyton floccosum (0.7%), Microsporum audouinii (1.1%), M. canis (3.1%), M. gypseum (0.3%), Trichophyton concentricum (3.5%), T. equinum (0.2%), T. mentagrophytes (36.1%), T. rubrum (53.8%), T. verrucosum (0.2) and T. violaceum (1.0%). The body sites most frequently affected by dermatophytes were the buttocks, nails and trunk. Anthropophilic dermatophytes made up 60.1% of the isolates; the most common species was T. rubrum, T. mentagrophytes and M. canis were the two main zoophilic dermatophytes. T. mentagrophytes was isolated from all body sites except the scalp. M. canis was found to be associated with domestic dogs and was not isolated from ethnic Malays. The only geophilic dermatophyte was M. gypseum, an uncommon dermatophyte associated with tinea pedis.     

西藏台错晚第四纪轮藻类的新类型

本文的研究材料来自西藏羌塘高原西部台错古湖一个3.69 m的天然剖面,其地层测年为32.6-4.5ka.从其156个样品中获取大量的轮藻类、介形类和腹足类及少量有孔虫等微体化石.据轮藻类藏卵器外形,顶部结构、螺旋细胞钙化式样等特征,该剖面的轮藻类应归属于轮藻科Characeae之轮藻属Chara.经研究得知,台错古湖在...     

Randomly amplified polymorphic DNA (RAPD) PCR for the identification of yeasts isolated from dairy products

In the present work randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) with primers M13 and RF2 was applied to the identification at species level of yeast strains isolated from cheeses. RAPD-PCR analysis of the type strains of different yeast species gave distinctive band profiles that allowed a clear differentiation of all the considered species. Forty-two of the 48 dairy associated yeasts were clearly assigned to the species Saccharomyces cerevisiae, Kluyveromyces marxianus (anamorph Candida kefyr), Kluyveromyces lactis (anamorph Candida sphaerica), Debaryomyces hansenii (anamorph Candida famata), Yarrowia lipolytica and Torulaspora delbrueckii (anamorph Candida colliculosa). The method, which is rapid and easy to perform, could be a useful tool for the identification of yeasts present in dairy products.     

Genus <Emphasis Type="Italic">Chelonus</Emphasis> Jurine (Hymenoptera,Braconidae, Cheloninae) in the fauna of Russia and adjacent territories. <Emphasis Type="Italic">Ch. olgae</Emphasis> Kok. species-group

The Ch. olgae species-group is established in the genus Chelonus characterized by the metasomal carapace narrowing towards the apex (much more strongly than towards the base) and not or weakly incurved. This group is analogous to the M. retusus group of the closely related genus Microchelonus, although some differences in the carapace structure emphasize parallel development of these morphological features in the two genera. New species are described as follows: Ch. borisi sp. n. (Russia: Leningrad Prov.), Ch. cardui sp. n. (Russia: Voronezh Prov.; Moldova), Ch. kokoujevi sp. n. (Russia: Belgorod and Chelyabinsk provinces), Ch. speijeri sp. n. (Spain), Ch. ubsunuricus sp. n. (Russia: Tuva), and Ch. yurii sp. n. (Kazakhstan). Chelonus subannulatus Abdinbekova, 1971 is synonymised with Ch. submuticus Wesmael, 1835(syn. n.).     

Cytochrome P-450 cholesterol 7 alpha-hydroxylase: inhibition of enzyme deactivation by structurally diverse calmodulin antagonists and phosphatase inhibitors

Cytochrome P-450 cholesterol 7 alpha-hydroxylase (P-450Ch7 alpha) catalyzes the first and rate-limiting step in the conversion of cholesterol to bile acids. Incubation of rat liver microsomes in 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid buffer resulted in a time-dependent deactivation of P-450Ch7 alpha which was markedly accelerated by the nonionic detergent Tween 80. Microsomal NADPH-cytochrome P-450 reductase and cytochrome P-450-dependent 7-ethoxycoumarin O-deethylase activities were unaffected under these conditions, evidencing the selectivity of the deactivation process for P-450Ch7 alpha. The rate (t 1/2 = 15-19 min at 37 degrees C) and maximal extent of P-450Ch7 alpha deactivation (greater than or equal to 90%) were both unaffected by the presence of cytosolic proteins and were also not dependent on the initial enzyme level, as shown using liver microsomes isolated from untreated, cholestyramine-fed, and xenobiotic-induced rats exhibiting an eight-fold range in P-450Ch7 alpha activity. Scavengers for reduced oxygen species were also without effect. P-450Ch7 alpha was stabilized some six- to sevenfold (t 1/2 = 94-143 min) by the phosphatase inhibitor NaF. Of a series of other phosphatase inhibitors examined, including, among others, EDTA, vanadate, and molybdate, only phosphate-containing compounds and the calmodulin antagonist trifluoperazine, and inhibitor of the Ca2+-calmodulin-dependent phosphatase calcineurin, effectively stabilized P-450Ch7 alpha. Modulation of P-450Ch7 alpha deactivation by these inhibitors generally paralleled their effects on isolated calcineurin. A variety of structurally diverse calmodulin antagonists examined were also found to effectively protect P-450Ch7 alpha from deactivation; these include calmidazolium and tamoxifen (IC50 = 25 to 50 microM), chlorpromazine, thioridazine, amitriptyline, imipramine, and the naphthalene sulfonamide compound W-7 (IC50 = 50 to 300 microM). Structure-activity analysis of several phenothiazines and their derivatives indicated that although little activity was exhibited by the sulfoxides, some protection was provided by the corresponding sulfones. On the basis of these observations, various models for the molecular basis of enzyme deactivation are considered, including the hypothesis that a calcineurin-like microsomal phosphatase mediates deactivation of this cytochrome P-450 enzyme.     

正常人耵聍中马拉色菌菌种构成研究

目的研究正常人耵聍中马拉色菌菌种构成及同一宿主耵聍中菌种是否一致。方法采集45名健康志愿者双侧耵聍,0.1%曲拉通X-100溶解稀释后接种于含菜籽油培养基,生化及形态学方法鉴定到种,同时提取菌种DNA,用真菌通用引物ITS1/ITS4做PCR扩增并测序鉴定。结果有44例(97.78%)双侧耵聍中均培养出马拉色菌(共分离出88株菌),菌种构成:糠粃马拉色菌29株(32.95%)、斯洛菲马拉色菌23株(26.14%)、合轴马拉色菌18株(20.45%)、球形马拉色菌11株(12.50%)、限制性马拉色菌7株(7.95%)。44例(88株菌)中双侧耵聍菌种相同者有38例(76株菌)(一致率86.36%)。结论正常人耵聍中马拉色菌菌种分布较广,主要菌种为糠秕马拉色菌。同一宿主双侧耵聍中马拉色菌菌种具有一定的一致性。     

Mycobacterium kumamotonense Sp. Nov. recovered from clinical specimen and the first isolation report of Mycobacterium arupense in Japan: Novel slowly growing, nonchromogenic clinical isolates related to Mycobacterium terrae complex

Three mycobacterium strains isolated from clinical specimens in Japan were provisionally assigned to the genus Mycobacterium based on their phenotypical characteristics. These isolates were further investigated to determine their specific taxonomic statuses. Mycolic acid analysis and 16S rRNA gene, rpoB, and hsp65 sequence data for the isolates showed that they are most similar to M. terrae complex. DNA-DNA hybridization studies indicated that the three strains were of two species and were distinguishable from M. terrae, M. nonchromogenicum, and M. hiberniae. Therefore, these strains represent two novel species within the genus Mycobacterium. However, one potential new species should have been considered as M. arupense with the 16S rRNA gene and hsp65 sequences similarities of 99.8% and 100% respectively; it was isolated from human specimens in the United States and was proposed in June 2006 as a new species. This report describes the first isolation of M. arupense in Japan, suggesting that the organism is clinically relevant. In addition, we propose the novel species designation Mycobacterium kumamotonense sp. nov. The type strain is CST 7247(T) (=GTC 2729(T), =JCM 13453(T), =CCUG 51961(T)).     

Multienzyme microbiosensor based on electropolymerized o-phenylenediamine for simultaneous in vitro determination of acetylcholine and choline

The electrochemical biosensors based on poly(o-phenylenediamine) (PoPD) and acetylcholinesterase (AChE) and choline oxidase (ChO) enzymes were fabricated on carbon fibre (CF) substrate. The electropolymerized PoPD was used to reduce the interfering substances. The electrode assembly was completed by depositing functionalized carbon nano tubes (FCNTs) and Nafion (Naf). Amperometric detection of acetylcholine (ACh) and choline (Ch) were realized at an applied potential of +750 mV vs Ag/AgCl (saturated KCl). At pH 7.4, the final assembly, Naf-FCNTs/AChE-ChO((10:1))/PoPD/CF(Elip), was observed to have high sensitivity towards Ch (6.3±0.3 μA mM(-1)) and ACh (5.8±0.3 μA mM(-1)), linear range for Ch (K(M)=0.52±0.03 mM) and ACh (K(M)=0.59±0.07 mM), and for Ch the highest ascorbic acid blocking capacity (97.2±2 1mM AA). It had a response time of <5s and with 0.045 μM limit of detection. Studies on different ratio (ACh/Ch) revealed that 10:1, gave best overall response.     

Copepod parasites of the fatheads (pisces, psychrolutidae) and their implication on the phylogenetic relationships of psychrolutid genera

Nine species of psychrolutids kept in the Marine Zoology of Hokkaido University were found to carry eight species of copepod parasites. The parasites and their hosts are: Bobkabata kabatabobbus Hogans and Benz, 1990 on Malacocottus zonurus Bean; Chondracanthus parvus n. sp. on Eurymen gyrinus Gilbert and Burke; Chondracanthus yabei n. sp. on Dasycottus setiger Bean and M. zonurus; Ch. yanezi Atria, 1980 on Psychrolutes phrictus Stein and Bond; Caligus similis n. sp. on Neophrynichthys latus (Hutton); Clavella adunca (Str?m, 1762) on M. zonurus; Neobrachiella amphipacifica Ho, 1982 on Ambophthalmos angustus (Nelson), Cottunculus sp., D. setiger, Ebinania brephocephala (Jordan and Starks), E. vermiculata Sakamoto, and P. phrictus; and Naobranchia occidentalis Wilson, 1915 on D. setiger and M. zonurus. Chondracanthus parvus is closest to Ch. deltoideus Fraser, 1920, but differs from it in having only one pair of small knobs on the head and carrying a pair of lateral processes on the second pediger. Chondracanthus yabei resembles Ch. yanezi Atria, 1980, but can be distinguished from it by the presence of three, low protrusions on the mid-dorsal surface of the trunk; besides, maxillule also shows difference. As the name indicates, C. similis resembles several species of Caligus that bear a short abdomen and a formula of I; IV on the exopod of leg 4. However, it can be separated from them by the combination of the following characters: a genital complex distinctly smaller than the cephalothoracic shield, a pair of relatively short caudal rami, a smooth dentiform process on maxillule, and a pair of truncate tines on sternal furca. Analysis of the occurrence of Chondracanthus on the psychrolutid fishes shows that the phylogeny of Chondracanthus is in congruence with that of the Psychrolutidae. It implies that parasitism of Chondracanthus occurred after the Dasycottus clade diverged off the main stock of the Psychrolutidae and the occurrence of Ch. yabei on D. setiger is resulted from a later colonization.     

High growth rate and regeneration capacity of hypocotyl protoplasts in some Brassicaceae

Protoplasts isolated from 4-day-old hypocotyls of various species of Brassica (Brassica napus, B. campestris and B. oleracea ) produced callus with high efficiency in media containing casein hydrolysate and high concentrations of the auxin 2,4-D (4.5 μM). Cell division began after 24 h and 60% of the cells had divided after 48 h. In contrast, protoplasts isolated from stem and mesophyll of plants grown in vitro or in the greenhouse began to divide after a delay of 3–5 days. In these cases 40–50% of the cells had divided after 5 days as compared to 70% for hypocotyl protoplasts. To obtain a high frequency of regeneration, rapidly growing calli were transferred to media having a high cytokinin:auxin ratio as early as possible, usually 3 weeks after protoplast isolation. The average regeneration frequency for calli obtained from mesophyll protoplasts was 50%, while as many as 70% of the calli derived from hypocotyl protoplasts of B. napus regenerated plantlets on a medium containing zeatin (9.1 μM) and IAA (0.6 μM). On the same medium regeneration of Brassica oleracea was obtained. A low percentage of calli (1%) from Brassica campestris formed shoots when cultured on a combination of zeatin (4.6 μM), BA (4.4 μM) and IAA (0.6 μM).     

Specific detection of the Old World screwworm fly,Chrysomya bezziana,in bulk fly trap catches using real‐time PCR

The Old World screwworm fly (OWS), Chrysomya bezziana Villeneuve (Diptera: Calliphoridae), is a myiasis‐causing blowfly of major concern for both animals and humans. Surveillance traps are used in several countries for early detection of incursions and to monitor control strategies. Examination of surveillance trap catches is time‐consuming and is complicated by the presence of morphologically similar flies that are difficult to differentiate from Ch. bezziana, especially when the condition of specimens is poor. A molecular‐based method to confirm or refute the presence of Ch. bezziana in trap catches would greatly simplify monitoring programmes. A species‐specific real‐time polymerase chain reaction (PCR) assay was designed to target the ribosomal DNA internal transcribed spacer 1 (rDNA ITS1) of Ch. bezziana. The assay uses both species‐specific primers and an OWS‐specific Taqman^® MGB probe. Specificity was confirmed against morphologically similar and related Chrysomya and Cochliomyia species. An optimal extraction protocol was developed to process trap catches of up to 1000 flies and the assay is sensitive enough to detect one Ch. bezziana in a sample of 1000 non‐target species. Blind testing of 29 trap catches from Australia and Malaysia detected Ch. bezziana with 100% accuracy. The probability of detecting OWS in a trap catch of 50 000 flies when the OWS population prevalence is low (one in 1000 flies) is 63.6% for one extraction. For three extractions (3000 flies), the probability of detection increases to 95.5%. The real‐time PCR assay, used in conjunction with morphology, will greatly increase screening capabilities in surveillance areas where OWS prevalence is low.