Food Consumption of the Free-Living Aquatic Nematode Pelodera chitwoodi

A Cartesian diver respirometer was used to measure O₂ uptake and respiratory quotients at 25 C. Respiratory quotients were about 0.70 in starved nematodes, and 0.80 in third-stage and adult nematodes that had fed on bacteria. The energy output as measured by O₂ uptake was inversely related to the concentration of bacteria in the medium, indicating reduction in feeding effort. Feeding bacteria to third-stage nematodes in divers quickly resulted in peak respiration rates averaging 6.4 nl O₂/μg wet weight nematode per hour (QO₂) or six times the endogenous rate. In about 4 hr, the rates fell and then stabilized at a QO₂ of 2.5. Adult males fed bacteria in divers had a peak QO₂ of 2.8 or twice the starved rate. Adult females fed bacteria had a peak QO₂ of 3.7. Starving adult males and third-stage larvae were estimated to lose 2.4% and 1.4%, respectively, of their body weight per day in the form of fat based on the caloric equivalent o f oxygen used and a respiratory quotient of 0.70. The caloric content of the bacteria fed to nematodes in divers was determined. It was then calculated that both third-stage larvae and adult males ingested bacteria equivalent to 4.4 × 10⁻⁵ cal/μg wet weight nematode tissue per hour when feeding. Of the bacterial calories ingested, the larvae used 27% and adults 21% for respiration. It was estimated that males ingested 3.1 × 10⁶ bacteria and females 10 × 10⁶ bacteria during an 8-day life span.     

Feather keratin hydrolysis by a Vibrio sp. strain kr2

The aim of the study was to characterize feather-degrading bacteria isolated from poultry industry waste. A Vibrio sp. strain kr2 producing a high keratinolytic activity when cultured on native feather-containing broth was isolated. The bacterium grew with an optimum at pH 6.0 and 30 degrees C, where maximum featherdegrading activity was also observed. Keratinase production was similar at both 25 and 30 degrees C, while the maximum concentration of soluble protein was reached at 30 degrees C. Reduction of disulphide bridges was also observed, increasing with cultivation time. The keratinase of strain kr2 was active on azokeratin, azocasein, benzoyl-arginine-p-nitroanilide and Ala-Ala-p-nitroanilide as substrates. The amino acid composition of the feather hydrolysate was determined, presenting similarities with that reported for feather lysate, feather meal and raw feathers. A novel feather-degrading bacterium was isolated and characterized, showing high keratinolytic activity. Complete feather degradation was achieved during cultivation. Strain kr2 shows potential for use for biotechnological processes involving keratin hydrolysis.     

Vibrio neonatus sp. nov. and Vibrio ezurae sp. nov. isolated from the gut of Japanese abalones

Five alginolytic, facultative anaerobic, non-motile bacteria were isolated from the gut of Japanese abalones (Haliotis discus discus, H. diversicolor diversicolor and H. diversicolor aquatilis). Phylogenetic analyses based on 16S rRNA gene and gap gene sequences indicated that these strains are closely related to V. halioticoli. DNA-DNA hybridizations, FAFLP fingerprintings, and phylogenies of gap and 16S rRNA gene sequences showed that the five strains represent two species different from all currently described vibrios. The names Vibrio neonatus sp. nov. (IAM 15060T = LMG 19973T = HDD3-1T; mol% G+C of DNA is 42.1-43.9), and Vibrio ezurae sp. nov. (IAM 15061T = LMG 19970T = HDS1-1T; mol% G+C of DNA is 43.6-44.8) are proposed to encompass these new taxa. The two new species can be differentiated from V. halioticoli on the basis of several features, including beta-galactosidase activity, assimilation of glycerol, D-mannose and D-gluconate.     

Unusual and Strongly Structured Sequence Variation in a Complex Satellite DNA Family from the Nematode Meloidogyne chitwoodi

An AluI satellite DNA family has been isolated in the genome of the root-knot nematode Meloidogyne chitwoodi. This repeated sequence was shown to be present at approximately 11,400 copies per haploid genome, and represents about 3.5% of the total genomic DNA. Nineteen monomers were cloned and sequenced. Their length ranged from 142 to 180 bp, and their A + T content was high (from 65.7 to 79.1%), with frequent runs of As and Ts. An unexpected heterogeneity in primary structure was observed between monomers, and multiple alignment analysis showed that the 19 repeats could be unambiguously clustered in six subfamilies. A consensus sequence has been deduced for each subfamily, within which the number of positions conserved is very high, ranging from 86.7% to 98.6%. Even though blocks of conserved regions could be observed, multiple alignment of the six consensus sequences did not enable the establishment of a general unambiguous consensus sequence. Screening of the six consensus sequences for evidence of internal repeated subunits revealed a 6-bp motif (AAATTT), present in both direct and inverted orientation. This motif was found up to nine times in the consensus sequences, also with the occurrence of degenerated subrepeats. Along with the meiotic parthenogenetic mode of reproduction of this nematode, such structural features may argue for the evolution of this satellite DNA family either (1) from a common ancestral sequence by amplification followed by mechanisms of sequence divergence, or (2) through independent mutations of the ancestral sequence in isolated amphimictic nematode populations and subsequent hybridization events. Overall, our results suggest the ancient origin of this satellite DNA family, and may reflect for M. chitwoodi a phylogenetic position close to the ancestral amphimictic forms of root-knot nematodes. Received: 23 April 1997 / Accepted: 9 July 1997     

Recovery from nutrient starvation by a marine Vibrio sp

A marine psychrophilic Vibrio sp., Ant-300, recovered from starvation after the addition of 1 volume of complete nutrient medium to 9 volumes of starvation menstruum. Turbidity (measured by optical density), viable cell counts, cell size (measured from electron micrographs), and cellular concentrations of protein, DNA, and RNA were monitored with recovery time. The usual growth curve of bacterial cultures was observed. On a per viable cell basis, protein, DNA, and RNA increased to maximum values just before cell division and then returned to close to the initial starved-cell value during the stationary phase. Cells under complete starvation conditions or missing only one nutrient in the stationary phase responded with cell division resulting in many smaller cells. The length of the lag phase during recovery was directly proportional to the length of the prior starvation period, even when identical numbers of cells were used for recovery. Cells appeared to pass more deeply into dormancy with starvation time.     

Meloidogyne megatyla n. sp. a Root-Knot Nematode from Loblolly Pine

Meloidogyne megatyla n. sp. is described from Pinus taeda in North Carolina. Stylet knobs are distinctively high in proportion to width, giving an especially massive appearance to the knobs of larvae and males. Mean larval length is 416 μm and stylet length is 14.6 μm. The perineal pattern is composed of smooth striae, with a high arch, and is often somewhat rectangular. The relationship of M. megatyla to other Meloidogyne species is unclear, although a comparison is made with Meloidogyne incognita and Meloidogyne mali. Galling was slight; only about 50 eggs were produced per egg mass, and under greenhouse conditions a single generation may take more than 10 weeks. Meloidogyne megatyla n. sp. did not reproduce on any of the differential hosts commonly used to distinguish among Meloidogyne species.     

Vibrio navarrensis sp. nov., a species from sewage.

A group of 11 strains, mostly isolated from sewage water in the Province of Navarra, Spain, were found to constitute a DNA relatedness group which is 2 to 39% related to 23 species of the genus Vibrio and 2 to 3% related to two Aeromonas species. Phenotypically, these strains have all of the properties that define the genus Vibrio. However, they differ from the previously described species by three or more properties. The strains are negative for arginine, ornithine, and lysine decarboxylase activities and the Voges-Proskauer test and are unable to utilize putrescine, gluconate, glucuronate, and histidine. They utilize and produce acid from sucrose and grow at 40 degrees C. All strains grow in the presence of 0.5% (wt/vol) NaCl, and seven strains grow weakly in peptone water lacking NaCl. The group of strains which we studied can also be differentiated from other Vibrio species by fatty acid content. The G+C ratio of the DNA is 45 to 47 mol%. The name Vibrio navarrensis sp. nov. is proposed for these strains; strain 1397-6 (= CIP 103381) is the type strain.     

Bionomics of a Phoretic Association Between Paenibacillus sp. and the Entomopathogenic Nematode Steinernema diaprepesi

Spores of an unidentified bacterium were discovered adhering to cuticles of third-stage infective juvenile (IJ) Steinernema diaprepesi endemic in a central Florida citrus orchard. The spores were cup-shaped, 5 to 6 mm in length, and contained a central endospore. Based on 16S rDNA gene sequencing, the bacterium is closely related to the insect pathogens Paenibacillus popilliae and P. lentimorbus. However, unlike the latter bacteria, the Paenibacillus sp. is non-fastidious and grew readily on several standard media. The bacterium did not attach to cuticles of several entomopathogenic or plant-parasitic nematodes tested, suggesting host specificity to S. diaprepesi. Attachment of Paenibacillus sp. to the third-stage cuticle of S. diaprepesi differed from Paenibacillus spp. associated with heterorhabditid entomopathogenic nematodes, which attach to the IJ sheath (second-stage cuticle). The inability to detect endospores within the body of S. diaprepesi indicates that the bacterial association with the nematode is phoretic. The Paenibacillus sp. showed limited virulence to Diaprepes abbreviatus, requiring inoculation of larvae with 108 spores to achieve death of the insect and reproduction of the bacterium. The effect of the bacterium on the nematode population biology was studied in 25-cm-long vertical sand columns. A single D. abbreviatus larva was confined below 15-cm depth, and the soil surface was inoculated with either spore-free or spore-encumbered IJ nematodes. After 7 days, the proportion of IJ below 5-cm depth was seven-fold greater for spore-free IJ than for spore-encumbered nematodes. Mortality of D. abbreviatus larvae was 72% greater (P <= 0.01) for spore-free compared to spore-encumbered S. diaprepesi. More than 5 times as many progeny IJs (P <= 0.01) were produced by spore-free compared to spore-encumbered nematodes. These data suggest that the bacterium is a component of the D. abbreviatus food web with some potential to regulate a natural enemy of the insect.     

Meloidogyne morocciensis n. sp. (Meloidogyninae), a Root-knot Nematode from Morocco

Meloidogyne morocciensis n. sp. is described from specimens parasitic on peach rootstock from Morocco. This species exhibits a combination of morphological characters similar to M. arenaria, M. incognita, and M. javanica. The perineal pattern of females is oval to squarish with a moderately high to high dorsal arch, and widely spaced, smooth striae; lateral lines are absent. The stylet, 16.5 μm long, has transversely ovoid, set-off knobs. Males have a set-off, annulated head region. The large, rounded labial disc is distinctly demarcated from the crescent-shaped medial lips; lateral lips are absent. The robust stylet, 24.6 μm long, has large, rounded knobs that taper slightly posteriorly. Mean second-stage juvenile (J2) length is 401 μm. The set-offhead region has incomplete annulations; the lip structures are dumbbell shaped. The stylet, 12.3 μm long, has rounded knobs that slope posteriorly. The J2 tail, 52.6 μm long, has irregularly sized annules in the posterior region and ends in a bluntly rounded tip. Tomato, tobacco, pepper, and watermelon are good hosts; cotton and peanut are not hosts. Meloidogyne morocciensis n. sp. reproduces by mitotic parthenogenesis and has a somatic chromosome number of 47-49. Its esterase phenotype is identical with the three-banded phenotype (A3) of M. arenaria.     

Novel transmissible factors in a non-O1 Vibrio cholerae and a Vibrio sp

Transmissible factors encoding production of lacunae (L factors) were demonstrated in a non-O1 Vibrio cholerae and a Vibrio sp. of recent environmental origin. Lacunae were produced in lawns of non-O1 V. cholerae indicator strains under the same assay conditions as those where lacunae were produced by the well characterized P fertility plasmid of V. cholerae O1 and the V fertility factor found in a non-cholera vibrio strain. The origin of the lacunae produced by strains harbouring the V and L factors was examined. No vibriocin or phage activity was found in culture supernates or in lacunae produced by the strains, suggesting that, as in the case of the P plasmid, the lacunae probably represent sites of active mating. Unlike the P plasmid, neither the Vn or L factor could be detected or isolated by conventional plasmid techniques.     

Dolichodorus aestuarius n. sp. (Nematode: Dolichodoridae)

Dolichodorus aestuarius n. sp. from an estuarine habitat near Cedar Key, Florida is described. This nematode has a stylet range of 62-76 μm in females and 60-72 μm in males. The stylet is shorter than those of all described species except D. brevistilus. The probable host plant is Juncus roemerianus.     

ATP level of a starving surface-bound and free-living marine Vibrio sp.

Abstract ATP levels of three marine isolates (a Vibrio sp., Serratia marcescens , and an unidentified rodshaped organism) were measured during nutrient-limiting conditions for free-living bacteria or cells adhered to a glass/liquid interface. In all strains there was a higher content of ATP per biovolume (numbers × cell volume) for cells starved at the glass/liquid interface compared with those in the bulk phase. Leakage from viable cells and the formation of a conditioning film are suggested only partly to explain the elevated levels of ATP per biovolume at the surface.     

Vibrio aestivus sp. nov. and Vibrio quintilis sp. nov., related to Marisflavi and Gazogenes clades,respectively

Two new Vibrio species, Vibrio aestivus and Vibrio quintilis, are described after a polyphasic characterization of strains M22^T, M61 and M62^T, isolated from seawater collected off a beach on the East coast of Spain (Valencia). All three strains are Gram negative, mesophilic, slightly halophilic, fermentative rods. V. aestivus (M22^T = CECT 7558^T = CAIM 1861^T = KCTC 23860^T and M61 = CECT 7559 = CAIM 1862 = KCTC 23861) is oxidase positive, reduces nitrates to nitrites, is negative for Voges Proskauer, arginine dihydrolase and indole and non hydrolytic on most substrates tested. The 16S rRNA gene sequences of M22^T and M61 are most similar to Vibrio marisflavi (97.1–97.2%) but phylogenetic analysis using NJ, MP and ML methods display Vibrio stylophorae (96.2% similarity) as sibling species. The three species form a deep clade in the genus Vibrio. Average Nucleotide Identity (ANI) values, determined as a measure of overall genomic resemblance, confirmed that strains M22^T and M61 are members of the same species, different to V. marisflavi CECT 7928^T.V. quintilis (M62^T = CECT 7734^T = CAIM 1863^T = KCTC 23833^T) is aerogenic, arginine dihydrolase and Voges Proskauer positive, oxidase negative and unable to reduce nitrate, traits shared by most species in the Gazogenes clade. It is unpigmented and does not grow on TCBS Agar. 16S rRNA gene similarities to its nearest species, Vibrio aerogenes and Vibrio mangrovi, are 97.6% and 96.0% respectively. Strain M62^T and V. aerogenes CECT 7868^T display ANI values well below the 95% boundary for genomic species.     

Isolation and characterization of chitinase from a marine bacterium Vibrio sp.

A chitinase was separated from the culture broth of Vibrio sp. 11211 isolated from sediment from the South China Sea. The chitinase was purified 18.3-fold with 33% recovery by ammonium sulphate precipitation and chromatography. The subunit molecular weight of the enzyme was estimated by SDS-PAGE to be about 30kDa. The enzyme showed optimum pH at 6.5 and optimum temperature at 50^°C, and was stable in the pH range of 4 to 9 and at the temperature below 40^°C.     

Lipid Composition of a Psychrophilic Marine Vibrio sp. During Starvation-Induced Morphogenesis

Qualitative and quantitative changes with time in phospholipids and fatty acids were examined after suspension of cells of a psychrophilic marine bacterium in nutrient-free artificial seawater at 5°C. Viability was maintained throughout the 21-day examination period, with plate counts and acridine orange direct counts indicating a slight increase in cell number. Gravimetric data, however, showed a significant decrease in bacterial biomass during the 3-week study. Levels of ATP per cell also decreased significantly (59%) during the starvation period. Since starvation (resulting in dormancy) is probably the typical physiological state of marine bacteria, estimation of bacterial density in marine waters by using ATP data obtained from log-phase cells is probably inaccurate. Total lipid phosphate decreased (65%) during the starvation period, with phosphatidylethanolamine showing the greatest loss. A large increase (57%) in the neutral lipid fraction was also detected, especially during the first week of starvation. A selective increase in palmitoleate at the expense of myristate was detected in the membrane lipids. The effects of these changes on membrane fluidity and the possible consequences for these cells in the marine environment are discussed.     

Production of feather protein hydrolysate by keratinolytic bacterium Vibrio sp. kr2

A feather protein hydrolysate was produced using the keratinolytic bacterium Vibrio sp. strain kr2. Complete feather degradation was observed in medium containing up to 60 g L(-1) raw feathers. Cultivation on 40, 60 or 80 g L(-1) feathers for five days resulted in similar amounts of soluble protein, reaching maximum values around 2.5 g L(-1). Maximum yields of soluble protein were achieved at 30 degrees C and initial pH ranging from 6.0 to 8.0. Strain kr2 was effective in producing keratin hydrolysate from chicken feathers. Bacterial feather hydrolysate has the potential for utilization as an ingredient in animal feed or as organic fertilizer, thereby reducing the environmental impact of feather waste from the poultry industry.     

Growth characteristics of an obligately psychrophilic Vibrio sp.

The growth characteristics of an obligately psychrophilic Vibrio sp. have been studied in a chemostat with glucose or lactose as the limiting substrate over a temperature range 0–23°C. Vibrio AF-1 has an optimum growth temperature of 15°C and maximum growth temperature which is dependent upon the carbon source. On glucose growth ceases at 20°C whereas on lactose growth continues to 23°C. Growth rate is also a function of the carbon source provided. When grown on glucose, fructose, sucrose, maltose and galactose _max values of 0.046 h^-1 at 15°C were recorded whereas on lactose, mannose, ribose and xylose _max values of 0.020 h^-1 were obtained. Substrate affinities (K _s ) for the 9 sugars also fall into 2 divisions as for _max and are temperature dependent. Those sugars which support a high growth rate have highest K _s values at 0°C whereas these which give a low growth rate show maximum affinities at 15°C. Vibrio AF-1 produces the maximum cell yield (0.6 g/g sugar consumed) at temperature <8°C irrespective of the carbon source utilised and correlated with maximum rates of sugar uptake and minimum O₂ consumption. Maintenance energy determination on glucose grown cells show that at 2° C 2% of the carbon input is used for maintenance whereas at 20°C the requirement increases to 10% of the carbon input.     

Protein Patterns of Growing and Starved Cells of a Marine Vibrio sp.

Fingerprint protein patterns were produced by two-dimensional polyacrylamide electrophoresis on lysed cells of a Vibrio sp., Ant-300, which were prepared from growing and starved cultures. The cells were labeled with [³⁵S]methionine during growth and subsequently starved for up to 30 days. Samples were taken at selected time points representing stages in the starvation-survival process. Unlabeled starved cells were allowed to recover in the presence of [³⁵S]methionine to determine protein changes associated with the recovery from starvation. All growth conditions produced similar protein fingerprints; however, some protein spots disappeared, whereas others were seen only during starvation.