Isolation of Naegleria australiensis from an Oklahoma Lake1

Eight isolates of Naegleria australiensis were obtained from a small lake in Tulsa, Oklahoma. The eight strains were isolated during the hot summer months of July through September, when water temperatures ranged from 27 to 33°C. All eight isolates were pathogenic for mice. The mean time to death for mice was 10 days (range 6–13 days). This pathogenic free-living ameba has not before been reported from the United States or the Western Hemisphere.     

Biochemical and biological characterization of ruminal Fusobacterium necrophorum

Abstract Biochemical characteristics, biological activities, and antimicrobial susceptibilities of ruminal Fusobacterium necrophorum (eight subsp. necrophorum and eight subsp. funduliforme ) and of isolates (three of each subsp.) obtained from bovine hepatic abscesses were determined. F. necrophorum subsp. necrophorum strains had higher phosphatase and DNase activities, produced more leukotoxin, and were more pathogenic to mice than subsp. funduliforme strains. The leukotoxin titer for culture supernatants of ruminal subsp. necrophorum strains was approximately 15 times lower than that of hepatic subsp. necrophorum strains. Hemagglutination activity was present in all hepatic, but only in some ruminal, strains of subsp. necrophorum . The antimicrobial sensitivity profile of the ruminal isolates was similar to that of hepatic isolates.     

Host Specificity of Pathogenic Pythium Species: Implications for Tree Species Diversity

In the Janzen–Connell hypothesis, host-specific natural enemies enhance species diversity and influence the structure of plant communities. This study tests the explicit assumption of host specificity for soil pathogens of the genus Pythium that cause damping-off disease of germinating seeds and seedlings. We isolated Pythium spp. from soil of a tropical forest in Panama. Then, in an inoculation experiment, we determined the pathogenicity of 75 tropical isolates of unknown pathogenicity and seven pathogenic temperate isolates of Pythium on seeds and/or seedlings of eight tropical tree species. Only three tropical isolates, one identified as P. ultimum and two as P. aphanidermatum , were pathogenic. Tropical pathogenic isolates were pathogenic on 4–6 of eight tree species. Temperate isolates were pathogenic on 0–4 of eight species, indicating that some tropical tree species are susceptible to novel isolates of Pythium . No tree species was susceptible to all isolates and two species were not susceptible to any isolate. Collectively, these results indicate that these Pythium isolates vary widely in their pathogenicity, causing differential mortality of potential host species; likewise, the tree species vary in their susceptibility to a given Pythium isolate. These differences in pathogenicity and susceptibility indicate some support for the Janzen–Connell assumption of host specificity. While they are not restricted to a single species, their intermediate level of specificity suggests that Pythium spp. have the potential to have some effect on forest community structure and diversity.     

Characterization of highly pathogenic H5N1 avian influenza A viruses isolated from South Korea

An unprecedented outbreak of H5N1 highly pathogenic avian influenza (HPAI) has been reported for poultry in eight different Asian countries, including South Korea, since December 2003. A phylogenetic analysis of the eight viral genes showed that the H5N1 poultry isolates from South Korea were of avian origin and contained the hemagglutinin and neuraminidase genes of the A/goose/Guangdong/1/96 (Gs/Gd) lineage. The current H5N1 strains in Asia, including the Korean isolates, share a gene constellation similar to that of the Penfold Park, Hong Kong, isolates from late 2002 and contain some molecular markers that seem to have been fixed in the Gs/Gd lineage virus since 2001. However, despite genetic similarities among recent H5N1 isolates, the topology of the phylogenetic tree clearly differentiates the Korean isolates from the Vietnamese and Thai isolates which have been reported to infect humans. A representative Korean isolate was inoculated into mice, with no mortality and no virus being isolated from the brain, although high titers of virus were observed in the lungs. The same isolate, however, caused systemic infections in chickens and quail and killed all of the birds within 2 and 4 days of intranasal inoculation, respectively. This isolate also replicated in multiple organs and tissues of ducks and caused some mortality. However, lower virus titers were observed in all corresponding tissues of ducks than in chicken and quail tissues, and the histological lesions were restricted to the respiratory tract. This study characterizes the molecular and biological properties of the H5N1 HPAI viruses from South Korea and emphasizes the need for comparative analyses of the H5N1 isolates from different countries to help elucidate the risk of a human pandemic from the strains of H5N1 HPAI currently circulating in Asia.     

Pathogenicity of Metarhizium anisopliae (Metsch.) Sorokin and Beauveria bassiana (Balsamo) Vuillemin,to three adult fruit fly species: Ceratitis capitata (Weidemann), C. rosa var. fasciventris Karsch and C. cosyra (Walker) (Diptera :Tephritidae)

The pathogenicity of two isolates of Beauveria bassiana and 12 of Metarhizium anisopliae towards adult fruit flies, Ceratitis capitata and Ceratitis rosa var. fasciventris was tested in the laboratory. Fruit flies were exposed to dry conidia evenly spread on velvet material covering the inner side of a cylindrical plastic tube. All isolates tested were pathogenic to both species of fruit flies. Mortality ranged from 7 to 100% in C. capitata and from 11.4 to 100% in C. rosa var. fasciventris at 4 days post-inoculation. Six isolates, M. anisopliae ICIPE 18, 20, 32, 40, 41 and 62, were highly pathogenic to both C. capitata and C. rosa var. fasciventris. The LT90 values of the most pathogenic isolates ranged between 3-4 days in both insects. Because of the difficulties in rearing C. cosyra, only the isolates that were highly pathogenic to both C. rosa var. fasciventris and C. capitata were tested against adult C. cosyra. They caused mortality of between 72-78% at 4 days post-inoculation. The LT90 values in all the isolates did not exceed 4 days. One of the most pathogenic isolates, M. anisopliae ICIPE 20, was evaluated against C. capitata and C. rosa var. fasciventris in cage experiments using three autoinoculators (maize cob, cheesecloth and Petri dish) in an autoinoculative device consisting of plastic mineral bottle. Mortality of between 70-93% was observed in flies of both species that were captured from the cages and held under laboratory conditions. These results indicate the possibility of fruit fly suppression with entomopathogenic fungi using an autoinoculative device.     

禽源性大肠杆菌O_1、O_2和O_(78)分离株外膜蛋白型的研究

对江苏省25个禽源性大肠杆菌O_1、O_2和O_(78)分离株的外膜蛋白型进行了测定。用改良的N-十二烷酰肌氨酸法提取其外膜蛋白,经SDS-PAGE电泳后,用考马斯亮蓝法进行染色。结果,8个O_1分离株,9个O_2分离株分属2个OMP型,其中的1个为二者所共有;而8个O_(78)分离株的OMP型也与该型相同。表明从江苏省分离到的禽源性大肠杆菌具有多样性的OMP型,而且这3个血清型的分离株中存在着共同的OMP型。     

Amoebae from antarctic soil and water.

Samples of soil and water were taken from the McMurdo Sound-Dry Valley region of Antarctica. Of the 70 samples cultured, 22 yielded amoebae capable of clonal growth at 30 degrees C. None of the isolates was pathogenic for mice. Acanthamoeba isolates appeared to show better survival potential than Naegleria isolates.     

A new bioassay method reveals pathogenicity of Metarhizium anisopliae and Beauveria bassiana against early stages of Capnodis tenebrionis (Coleoptera; Buprestidae)

We used a newly developed bioassay method to demonstrate for the first time the potential of the entomopathogenic fungi Beauveria bassiana and Metarhizium anisopliae to be used for the control of neonate larvae of Capnodis tenebrionis, a major threat to stone-fruit orchards in several countries. Four B. bassiana and four M. anisopliae isolates were all pathogenic for neonate larvae of C. tenebrionis; mortality rates 10 days after inoculation by dipping in a suspension with 10(8)conidia/ml varied from 23.5% to 100%. Three of the four M. anisopliae isolates caused 100% mortality. In most cases, postmortem hyphal growth and sporulation of M. anisopliae or B. bassiana was observed covering the larvae in their galleries. The eight isolates were also evaluated for pathogenicity to C. tenebrionis eggs at the same dosage. Only two B. bassiana isolates caused significant egg hatching reduction of 84.5% and 94.5%. Our results indicate that M. anisopliae and B. bassiana may be considered as promising for a new approach to prevent larval infestations by C. tenebrionis.     

Virulence and Vegetative Compatibility of Dutch and Italian Isolates of Fusarium oxysporum f. sp. lilii

The virulence and vegetative compatibility of eight Dutch and four Italian isolates of Fusarium oxysporum obtained from lily were compared. The virulence was tested by determination of the specific interaction between the Fusarium isolates and eight lily cultivars. A specific interaction was not found, so the existence of races was not demonstrated. Six of the twelve isolates turned out to be non-pathogenic for lily. The pathogenic isolates fell in four vegetative compatibility groups. No vegetative compatibility was found between isolates of F. oxysporum f. sp. lilii and those of f. sp. gladioli.     

Occurrence and characterization of Acanthamoeba similar to genotypes T4, T5, and T2/T6 isolated from environmental sources in Brasília, Federal District, Brazil

Species of Acanthamoeba can cause keratitis and brain infections. The characterization of environmental isolates is necessary to analyze the risk of human infection. We aimed at identifying and genotyping Acanthamoeba isolates from soil, swimming pools, and water features in Brasília, Federal District, Brazil, as well as determining their physiological characteristics and pathogenic potential. Among the 18 isolates studied, eight were similar to genotype T5, five to T4, and one to T2/T6, classified by the sequence analysis of 18S rDNA. Genotypes of four isolates were not determined. Ten isolates (55%) grew at 37 °C and seven (39%) grew in media with 1.5M mannitol, which are the physiological parameters associated with pathogenic Acanthamoeba; also, four isolates from swimming pools presented high pathogenic potential. Our results indicate a widespread distribution of potentially pathogenic Acanthamoeba T4, T5, and T2/T6 in different environmental sources in Brasília, revealing the potential risk of human infection and the need of preventive measures.     

Genotyping and Antifungal Drug Susceptibility of Trichosporon asahii Isolated from Chinese Patients

As a major pathogenic agent of trichosporonosis, Trichosporon asahii can cause life-threatening infection in immunocompromised patients. In this study, we analyzed the genotypes of the intergenic spacer (IGS) 1 region of the rRNA gene and the antifungal drug susceptibility of eight T. asahii isolates obtained from Chinese patients. Five genotypes were identified from the eight isolates, including three novel genotypes, three genotype 1, and two genotype 4. The eight T. asahii isolates were resistant to amphotericin B, 5-flucytosine, and terbinafine, but were highly sensitive to fluconazole (FLC), itraconazole (ITC), and voriconazole (VRC). The mean minimum inhibitory concentrations (MICs) of FLC and VRC were significantly lower than those reported in most other countries, while that of ITC was slightly higher. Our results suggest that genotypes of the T. asahii isolated from China are different from those of other countries, and azole drugs appeared to be more effective on the Chinese isolates. These results provide new insights into the epidemiology and antifungal treatment for T. asahii.     

Characterization of Vibrio fluvialis-like strains implicated in limp lobster disease

Studies were undertaken to characterize and determine the pathogenic mechanisms involved in a newly described systemic disease in Homarus americanus (American lobster) caused by a Vibrio fluvialis-like microorganism. Nineteen isolates were obtained from eight of nine lobsters sampled. Biochemically, the isolates resembled V. fluvialis, and the isolates grew optimally at 20 degrees C; none could grow at temperatures above 23 degrees C. The type strain (1AMA) displayed a thermal reduction time (D value) of 5.77 min at 37 degrees C. All of the isolates required at least 1% NaCl for growth. Collectively, the data suggest that these isolates may embody a new biotype. Pulsed-field gel electrophoresis (PFGE) analysis of the isolates revealed five closely related subgroups. Some isolates produced a sheep hemagglutinin that was neither an outer membrane protein nor a metalloprotease. Several isolates possessed capsules. The isolates were highly susceptible to a variety of antibiotics tested. However, six isolates were resistant to erythromycin. Seventeen isolates harbored plasmids. Lobster challenge studies revealed that the 50% lethal dose of a plasmid-positive strain was 100-fold lower than that of a plasmid-negative strain, suggesting that the plasmid may enhance the pathogenicity of these microorganisms in lobsters. Microorganisms that were recovered from experimentally infected lobsters exhibited biochemical and PFGE profiles that were indistinguishable from those of the challenge strain. Tissue affinity studies demonstrated that the challenge microorganisms accumulated in heart and midgut tissues as well as in the hemolymph. Culture supernatants and polymyxin B lysates of the strains caused elongation of CHO cells in tissue culture, suggesting the presence of a hitherto unknown enterotoxin. Both plasmid-positive and plasmid-negative strains caused significant dose-related intestinal fluid accumulations in suckling mice. Absence of viable organisms in the intestinal contents of mice suggests that these microorganisms cause diarrhea in mice by intoxication rather than by an infectious process. Further, these results support the thermal reduction data at 37 degrees C and suggest that the mechanism(s) that led to fluid accumulation in mice differs from the disease process observed in lobsters by requiring neither the persistence of viable microorganisms nor the presence of plasmids. In summary, results of lobster studies satisfy Koch's postulates at the organismal and molecular levels; the findings support the hypothesis that these V. fluvialis-like organisms were responsible for the originally described systemic disease, which is now called limp lobster disease.     

Importance of flagella and enterotoxins for Aeromonas virulence in a mouse model

A genetic characterization of eight virulence factor genes, elastase, lipase, polar flagella (flaA/flaB, flaG), lateral flagella (lafA), and the enterotoxins alt, act, and ast, was performed using polymerase chain reaction with 55 drinking water and nine clinical isolates. When 16 Aeromonas hydrophila strains, seven Aeromonas veronii strains, and seven Aeromonas caviae strains exhibiting different combinations of virulence factor genes were tested in immunocompromised mice by intraperitoneal injection, only those strains that had one or more of the enterotoxins flaA, flaB, and either flaG or lafA showed signs of being virulent. The correlation was seen in 97% (29/30) of the strains, which included strains from drinking water. Thus, Aeromonas water isolates have the potential to be pathogenic in immunocompromised hosts.     

Pathogenicity of vibrio splendidus strains associated with turbot larvae, scophthalmus maximus

Turbot larvae were challenged with eight strains of Vibrio splendidus isolated from diseased larvae, plus a ninth strain pathogenic to scallop larvae (A515; Nicolas et al . 1996 ). Six strains caused heavy mortality but the scallop pathogen and the other two strains did not. All the strains shared a large number of phenotypic traits, and an attempt was made to relate virulence to genotype and phenotype. Five of the six pathogenic strains were very similar, as shown by RAPD fingerprinting and phenotypic characteristics. The relatedness of the other strains was intermediate between the main pathogenic group and V. splendidus ATCC 33125, but the DNA–DNA homology between the pathogenic group and the reference strain was still high (78% of reassociation rate). The non-pathogenic isolates may be a useful tool for determining the possible virulence factors, as all the isolates differed by few characteristics.     

The gamma interferon knockout mouse model for sarcocystis neurona: comparison of infectivity of sporocysts and merozoites and routes of inoculation.

The dose-related infectivity of Sarcocystis neurona sporocysts and merozoites of 2 recent isolates of S. neurona was compared in gamma interferon knockout (KO) mice. Tenfold dilutions of sporocysts or merozoites were bioassayed in mice, cell culture, or both. All 8 mice, fed 1,000 sporocysts, developed neurological signs with demonstrable S. neurona in their tissues. Of 24 mice fed low numbers of sporocysts (100, 10, 1), 18 became ill by 4 wk postinoculation, and S. neurona was demonstrated in their brains; antibodies (S. neurona agglutination test) to S. neurona and S. neurona parasites were not found in tissues of the 6 mice that were fed sporocysts and survived for >39 days. One thousand culture-derived merozoites of these 2 isolates were pathogenic to all 8 mice inoculated subcutaneously (s.c.). Of the 24 mice inoculated s.c. with merozoites numbering 100, 10, or 1, only 3 mice had demonstrable S. neurona infection; antibodies to S. neurona were not found in the 21 mice that had no demonstrable organisms. As few as 10 merozoites were infective for cell cultures. These results demonstrate that at least 1,000 merozoites are needed to cause disease in KO mice. Sarcocystis neurona sporocysts were infective to mice by the s.c. route.     

Lack of Toxicity of Helminthosporium maydis-Invaded Corn and Culture Filtrates to Chicks and Mice

Six isolates of Helminthosporium maydis, obtained from southern leaf blight-damaged corn, were grown separately on autoclaved corn and fed to chicks and mice to evaluate their toxigenicity. Two-, three-, and four-week-old culture filtrates from three pathogenic isolates grown separately on modified Fries medium were also evaluated for toxigenicity. None of the invaded corn samples or culture filtrates affected the weights of chicks or mice when compared to controls. Postmortem examinations did not reveal significant gross lesions. H. maydis was not toxigenic under our experimental conditions.     

Occurrence and characterization of "avium-like" Mycobacteria isolated from animals in Sweden

431 cultures of "avium-like" mycobacteria (ALM) were isolated from wild and domestic animals during 1974--76 at the National Veterinary Institute, Stockholm. Of these, 50 isolates from pigs were studied by growth-chromogenicity, pathogenicity, and biochemical tests. Furthermore, thin-layer chromathography was performed, and on some isolates serotyping. All 50 isolates belonged to Runyon's group III and were pathogenic for chicken; none was capable of splitting oleic acid from Tween 80. 47 gave tellurite reduction within a period of three days; one was arylsulphatase-positive after three days and a further four after 14 days. The biological and biochemical tests permit assignation of the 50 isolates to the M. avium-intracellulare complex. The lipid patterns of the isolates examined were analysed by thin-layer chromatography. Thirty-five of the isolates showed a lipid pattern similar to that of A 2 of the fowl reference strain; three belonged to lipid type A 1 and four to A 3. Eight could not be typed. Of 22 isolates, 14 could be assigned to M. avium serotypes.     

Ord River arboviruses--isolations from mosquitoes.

One hundred and thirty presumptive viruses have been isolated from 485 pools made from 23,872 mosquitoes collected in the Ord River area of North-West Australia. One hundred and eleven of the virus isolates isolates came from pools of Culex annulirostis, the dominant mosquito species caught in the vicinity of Kununurra. Forty-five of the viruses pathogenic for newborn mice have been further characterized-19 as Flaviviruses, 1 Alphavirus, 9 Koongol, 1 Mapputta and 15 non-haemagglutinating viruses of which 6 are Corripata. Thirty-seven isolates were from Culex annulirostis, 7 from Aedomyia catasticta and 1 from aedes tremulus. All Corriparta isolates were from Aedomyia catasticta. The Flaviviruses comprised 13 Kunjin and 6 MVE isolates.