Mycoflora isolation and molecular characterization of Aspergillus and Fusarium species in Tunisian cereals

Wheat, barley and maize are the mainly consumed cereals in Tunisia. This study aimed to determine the mycoflora of these cereals with special focus on the mycotoxigenic Aspergillus and Fusarium species. Freshly harvested samples and other stored samples of each type of cereal (31 and 34 samples, respectively) were collected in Tunisia and cultured for fungal isolation and identification. Identification of fungal genera was based on morphological features. Aspergillus and Fusarium species were identified by species specific PCR assays complemented with DNA sequencing. Alternaria (70.83%), Eurotium (62.50%), Aspergillus (54.17%) and Penicillium (41.67%) were the most frequent fungi isolated from wheat. Penicillium (75%), Aspergillus (70%), Eurotium (65%) and Alternaria (65%) were the most frequently recovered genera from barley. The predominant genera in maize were Aspergillus (76.19%), Eurotium (42.86%), and Penicillium (38.09%). Aspergilllus, Penicillium, Fusarium and Alternaria were detected in both stored and freshly harvested grain samples. The frequencies of contamination with Aspergillus, Fusarium and Alternaria were higher in freshly harvested samples, whereas Penicillium species were more frequent in stored samples. The predominant Aspergillus species detected were A. flavus and A. niger. The Fusarium species detected were F. equiseti, F. verticillioides, F. nygamai, and F. oxysporum. This study suggested the potential risk for Aflatoxins and, to a lesser extent, for Ochratoxin A in Tunisian cereals. This is the first survey about mycoflora associated with wheat, barley and maize in Tunisia.     

Fishborne Trematode Metacercariae in Freshwater Fish from Guangxi Zhuang Autonomous Region, China

A survey was performed to investigate the infection status of fishborne trematode (FBT) metacercariae in freshwater fish from Guangxi Zhuang Autonomous Region, China. A total of 307 freshwater fish of 31 species were collected from 5 administrative regions of Guangxi Zhuang Autonomous Region. They were examined by artificial digestion method from July 2003 to August 2004. No metacercariae were detected in fish from Fusui-xian. In fish from Mashan-xian and a market in Nanning, 3 species of metacercariae, Haplorchis taichui, Haplorchis pumilio, and Centrocestus formosanus, were mainly detected. Metacercariae (8 in number) of Clonorchis sinensis were found in 1 Chanodichthys dabryi purchased from a market in Nanning. In fish from Yangshuo, Metagonimus yokogawai metacercariae were detected from all 18 fish species examined. Total 13 C. sinensis metacercariae were found in 3 out of 10 Hemibarbus maculatus from Yangshuo. All 7 Zacco platypus from Yangshuo were infected with 8-112 Echinochasmus perfoliatus metacercariae. In fish from Binyang-xian, H. pumilo metacercariae were mainly detected in all 5 fish species examined, and only 1 metacercaria of C. sinensis was found in a Hemiculter leucisculus. From the above results, it was confirmed that some species of freshwater fish play a role of second intermediate hosts for FBT in Guangxi Zhuang Autonomous Region, China. In particular, 4 species of intestinal flukes, M. yokogawai, H. taichui, H. pumilio, and C. formosanus, were prevalent in fish hosts, whereas C. sinensis metacercariae were detected only in 3 fish species.     

Fishborne Trematode Metacercariae Detected in Freshwater Fish from Vientiane Municipality and Savannakhet Province, Lao PDR

Freshwater fish from Vientiane Municipality and Savannakhet Province, Lao PDR were examined by the muscle compression and artificial digestion methods to know the infection status with trematode metacercariae. In the fish from Savannakhet, 2 species of metacercariae, Opisthorchis viverrini and Haplorchis taichui, were detected. O. viverrini metacercariae were found in 6 species of fish, Puntius brevis, Hampala dispar, Esomus metallicus, Mystacoleucus marginatus, Puntioplites falcifer, and Cyclocheilichthys armatus. H. taichui metacercariae were detected in 3 species of fish, P. brevis, P. falcifer, and M. marginatus. In the fish from Vientiane, 4 species of metacercariae, O. viverrini, H. taichui, Haplorchis yokogawai, and Centrocestus formosanus, were detected. Among them, O. viverrini metacercariae were found in 7 species of fish, Onychostoma elongatum, C. armatus, H. dispar, P. brevis, Cyclocheilichthys repasson, Osteochilus hasseltii, and Hypsibarbus lagleri. The metacercariae of H. taichui were detected in 6 species of fish, C. repasson, O. elongatum, C. armatus, H. dispar, Labiobarbus leptocheila, and Cirrhinus molitorella. The metacercariae of H. yokogawai were found in 9 species of fish, C. repasson, O. elongatum, C. armatus, H. dispar, Labiobarbus leptocheila, O. hasseltii. C. molitorella, Hypsibarbus wetmorei, and H. lagleri. The metacercariae of C. formosanus were detected in 4 species of fish, C. repasson, P. brevis, O. hasseltii, and C. molitorella. From these results, it is confirmed that fishborne trematode metacercariae, i.e. O. viverrini, H. taichui, H. yokogawai and C. formosanus, are prevalent in various species of freshwater fish from Savannakhet Province and Vientiane Municipality, Lao PDR.     

Antagonists of Plant-parasitic Nematodes in Florida Citrus

In a survey of antagonists of nematodes in 27 citrus groves, each with a history of Tylenchulus semipenetrans infestation, and 17 noncitrus habitats in Florida, approximately 24 species of microbial antagonists capable of attacking vermiform stages of Radopholus citrophilus were recovered. Eleven of these microbes and a species of Pasteuria also were observed attacking vermiform stages of T. semipenetrans. Verticillium chlamydosporium, Paecilomyces lilacinus, P. marquandii, Streptomyces sp., Arthrobotrys oligospora, and Dactylella ellipsospora were found infecting T. semipenetrans egg masses. Two species of nematophagous amoebae, five species of predatory nematodes, and 29 species of nematophagous arthropods also were detected. Nematode-trapping fungi and nematophagous arthropods were common inhabitants of citrus groves with a history of citrus nematode infestation; however, obligate parasites of nematodes were rare.     

Klebsiella Biotypes Among Coliforms Isolated from Forest Environments and Farm Produce

Samples of water, soil, needles, and bark were collected from three different forest environments and from a pulp and paper mill. In addition, samples of fresh produce were obtained from a local supermarket. All samples were examined for total and fecal coliforms. The counts obtained from the forestrelated samples did not correlate with sample type or location. When 123 isolates were identified biochemically, 71% were Klebsiella, 19% Enterobacter, 8% Citrobacter, and 2% Escherichia. All the Citrobacter, 75% of the Enterobacter, and 65% of the Klebsiella were negative for growth in elevated coliform (EC) broth. All the Escherichia were EC positive. The counts obtained from the fresh produce were generally higher than the forest counts, but the distribution of biotypes was similar. Of the 146 isolates examined 64% were Klebsiella, 14% were Escherichia, 14% were Enterobacter, and 8% were Citrobacter. All the Enterobacter and Citrobacter were EC negative, whereas 25% of the Klebesiella and 80% of the Escherichia were EC positive.     

A polyphasic approach for the characterization of endophytic Alternaria strains isolated from grapevines

A polyphasic approach was set up and applied to characterize 20 fungal endophytes belonging to the genus Alternaria, recovered from grapevine in different Italian regions.Morphological, microscopical, molecular and chemical investigations were performed and the obtained results were combined in a pooled cluster analysis. Following morphological analyses, all strains were grouped according to their three-dimensional sporulation pattern on PCA and to the colony characteristics on different substrates. After DNA extraction, all strains were analyzed by RAPD-PCR and the resulting profiles were subjected to cluster analysis. The metabolites extracted from the 20 Alternaria endophytes were analyzed by a HPLC and the resulting metabolite profiles were subjected to multivariate statistic analyses. In comparison with reference ‘small-spored’ Alternaria species, the 20 strains were segregated into two morphological groups: one belonging to the A. arborescens species-group and a second to the A. tenuissima species-group. RAPD analysis also showed that grapevine endophytes belonged to either the A. arborescens or the A. tenuissima species-group and that they were molecularly distinct from strains belonging to A. alternata. Chemotaxonomy gave the same grouping: the grapevine endophytic strains belong to A. arborescens or A. tenuissima species-groups producing known metabolites typical of these species-groups. Interestingly, the 20 grapevine endophytes were able to produce also a number of unknown metabolites, whose characterization could be useful for a more precise segregation of the two species-groups.The results show how complementary morphological, molecular and chemical data can clarify relationships among endophyte species-groups of low morphological divergence.     

Nitrous oxide reductase (nosZ) gene-specific PCR primers for detection of denitrifiers and three nosZ genes from marine sediments

Two PCR primer sets for the nitrous oxide reductase gene (nosZ) were developed. The initial primers were based on three sequences in GenBank and used to amplify nosZ from continental shelf sediments and from two denitrifiers in culture, Thiosphaera pantotropha and Pseudomonas denitrificans. Three unique marine sediment nosZ genes were identified and sequenced. The marine nosZ genes were most closely related to the nosZ genes of Paracoccus denitrificans or to Rhizobium meliloti. Alignment of all nosZ sequences currently available (n=10) facilitated redesign of the PCR primers. Three new primer sets which amplify 1100 bp, 900 bp and 250 bp regions of the nosZ gene were designed and tested. The new primers robustly amplified nosZ fragments from samples in which the initial nosZ primers were only marginally successful.     

External and internal flavonoids from Madagascarian Uncarina species (Pedaliaceae)

External and internal flavonoids were isolated from 12 Uncarina taxa (Pedaliaceae), endemic to Madagascar. Four flavone aglycones, tricetin 7,3′,5′-trimethyl ether, tricetin 7,4′,5′-trimethyl ether, 5,3′-dihydroxy-6,7,4′,5′-tetramethoxyflavone and eupatorin were isolated from leaf wax of seven Uncarina taxa, Uncarina grandidieri, Uncarina decaryi, Uncarina abbreviata, Uncarina turicana, Uncarina platycarpa, Uncarina leandrii var. leandrii and Uncarina peltata, but not Uncarina stellulifera, Uncarina perrieri, Uncarina sakalava, Uncarina leptocarpa and U. leandrii var. rechbergeri. Furthermore, eight flavonoid glycosides were isolated from the leaves. Major glycosides were apigenin and luteolin 7-O-glucuronides and occurred in all the Uncarina taxa examined, except the absence of the former compound in U. peltata. Other glycosides were identified as hispidulin, jaceosidin, chrysoeriol and tricin 7-O-glucuronides, and luteolin 7,4′-di-O-glucuronide and a flavonol, isorhamnetin 3-O-diglucoside. From the results described above, methylated flavone aglycones and glucuronides were chemical characters of the leaves of Uncarina species, and also may be those of the family Pedaliaceae. Besides, an anthocyanin, two flavonols and three flavones were isolated from the flowers of U. grandidieri, and identified as cyanidin 3-O-rutinoside (anthocyanin), quercetin and isorhamnetin 7-O-glucuronides (flavonols) and apigenin, luteolin and jaceosidin 7-O-glucuronides (flavones).     

The roots of the halophyte Salicornia brachiata are a source of new halotolerant diazotrophic bacteria with plant growth-promoting potential

Soil salinity is the major cause limiting plant productivity worldwide. Nitrogen-fixing bacteria were enriched and characterised from roots of Salicornia brachiata, an extreme halophyte which has substantial economic value as a bioresource of diverse and valuable products. Nitrogen-free semisolid NFb medium with malate as carbon source and up to 4% NaCl were used for enrichment and isolation of diazotrophic bacteria. The isolates were tested for plant growth-promoting traits and 16S rRNA, nifH and acdS genes were analysed. For selected strains, plant growth-promoting activities were tested in axenically grown Salicornia seedlings at different NaCl concentrations (0–0.5M). New halotolerant diazotrophic bacteria were isolated from roots of S. brachiata. The isolates were identified as Brachybacterium saurashtrense sp. nov., Zhihengliuella sp., Brevibacterium casei, Haererehalobacter sp., Halomonas sp., Vibrio sp., Cronobacter sakazakii, Pseudomonas spp., Rhizobium radiobacter, and Mesorhizobium sp. Nitrogen fixation as well as plant growth-promoting traits such as indole acetic acid (IAA) production, phosphate solubilisation, and 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase activity were demonstrated. For Brachybacterium saurashtrense and Pseudomonas sp., significant plant growth-promoting activities were observed in Salicornia in salt stress conditions. Salicornia brachiata is a useful source of new halotolerant diazotrophic bacteria with plant growth-promoting potential.     

Distribution and diversity of rhizobia associated with wild soybean (Glycine soja Sieb. & Zucc.) in Northwest China

A total of 155 nodule isolates that originated from seven sites in Northwest China were characterized by PCR-RFLP of the 16S rRNA gene and sequence analysis of multiple core genes (16S rRNA, recA, atpD, and glnII) in order to investigate the diversity and biogeography of Glycine soja-nodulating rhizobia. Among the isolates, 80 were Ensifer fredii, 19 were Ensifer morelense, 49 were Rhizobium radiobacter, and 7 were putative novel Rhizobium species. The phylogenies of E. fredii and E. morelense isolates in a concatenate tree (assembly of all housekeeping genes) were generally consistent with those in individual gene trees. However, incongruence was found in the phylogenies of the different genes of Rhizobium isolates, indicating that lateral transfer or recombination possibly occurred in these gene loci. Despite their species identity, all the isolates in this study formed a single lineage related to E. fredii in nodAand nifH gene phylogenies, which also indicated that the symbiotic genes were laterally transferred between different species. Biogeographic patterns were found at the species and strain genomic type levels, as revealed by BOXA1R fingerprinting, demonstrating that the evolution of rhizobial populations in different geographic locations was related to soil types, altitude and spatial effects. This study is the first to report that E. morelense, R. radiobacter, and Rhizobium sp. are microsymbionts of G. soja, as well as showing that the diversity of G. soja rhizobia is enhanced and new rhizobia have evolved in Northwest China.     

Symbiotic microbial studies in diverse populations of Aphis gossypii,existing on altered host plants in different localities during different times

Complex interactions between symbiotic bacteria and insects ultimately result in equilibrium in all aspects of life in natural insect populations. In this study, abundance of principal symbiotic bacteria was estimated using qPCR in 1553 individuals of aphids, Aphis gossypii. Aphids were sampled from primary and secondary host plants—hibiscus and cotton. Hibiscus aphids were collected from 24 different locations in April, September, and November, whereas cotton aphids were collected between 2015 and 2017 from areas with wide variations in climatic conditions. About 30%–45% aphids were recorded with the most dominant symbiont, Arsenophonus. The other symbionts were in low frequency, and about 7% of aphids were noted with Hamiltonella, Acinetobacter, and Microbacterium, and 3% of aphids were verified with Serratia and Pseudomonas. Aphids infected with Hamiltonella, Arsenophonus, and Serratia can influence Buchnera densities. Hamiltonella has positive interaction with densities of Arsenophonus and Serratia. Almost 100% coinfection of Hamiltonella and Arsenophonus was detected in Xinxiang aphids and 50% coinfection was reported in aphids from North China, while no coinfection was detected in Hainan aphids. These findings describe the prevalence pattern and richness of core community of symbiotic bacteria in naturally occurring populations of A. gossypii and provide new insights for the study of symbiotic bacteria.     

Leishmania (Sauroleishmania) tarentolae isolation and sympatric occurrence with Leishmania (Leishmania) infantum in geckoes,dogs and sand flies

The trypanosomatid protist Leishmania tarentolae is a saurian-associated parasite vectored by the Sergentomyia minuta sand fly. This study aimed to confirm the circulation of L. infantum and L. tarentolae in sand flies, reptiles and dogs and to isolate new strains of these protists. Reptilian and sheltered dog blood samples were collected, and sand flies were captured. Samples were tested for Leishmania spp. using duplex real-time PCR (dqPCR) and real-time PCR (qPCR); the origin of blood meal was identified in engorged sand flies by conventional PCR. The reptilian blood and intestinal content of sand fly females were cultured. Dog sera were tested by IFAT using both Leishmania species. Four Tarentola mauritanica geckoes were molecularly positive for L. infantum or L. tarentolae, with no co-infections; moreover, amastigote-like forms of L. infantum were observed in the bone marrow. 24/294 sand flies scored positive for Leishmania spp. by dqPCR, 21 S. minuta and two Phlebotomus perniciosus were positive for L. tarentolae, while only a single Ph. perniciosus was positive for L. infantum. Blood meal analysis confirmed reptile and dog in S. minuta, dog and human in Ph. perniciosus and dog in Phlebotomus neglectus. Two axenic strains of L. tarentolae were obtained. Twelve of 19 dogs scored positive for L. infantum and L. tarentolae by IFAT and three of them also for L. infantum by dqPCR, and six by qPCR. These data confirm the sympatric circulation of L. infantum and L. tarentolae in geckoes, sand flies, and dogs, and suggest that geckoes may be infected with L. infantum.     

Identification and characterization of four new tra cistrons on the E. coli K12 sex factor F.

Thirty-eight amber-suppressible transfer-deficient mutants of the Flac element F13-1 have been analyzed in conjugational complementation tests. Many of the mutations were in tra genes that have been identified previously (including traI), but several were in four new genes, traN, traU, traV, and traW. Results for representative mutations in the new genes were confirmed by complementation tests using λtra transducing phages and these also allowed the genes to be mapped quickly and simply. Approximate physical locations were found by complementation with a series of chimeric plasmids carrying various EcoRI fragments of the F transfer region. All four genes are located in the transfer operon, the order being traA … traBtraVtraWtraCtraUtraNtraF … traD. While traU, traV, and traW mutants were resistant to male-specific phages, traN mutants were sensitive.     

The interaction of Babesia caballi kinetes with tick cells

Babesia caballi caused pathological changes in the vector tick, Anocentor nitents. These included the loss of hemocytes, reddish-brown discoloration of the hemolymph, retardation in clotting, and reduction in egg production. Primary cultures were initiated from A. nitens embryos transovarially infected with B. caballi. Cell lines that were isolated were morphologically similar and composed of stellate, fusiform, and hemocyte-like cells, but none was visibly infected with B. caballi. Primary cultures from heavily infected ticks were not viable and did not develop into lines. Kinetes from hemolymph and organs of A. nitens were cocultivated with cell lines isolated from embryos of A. nitens, Rhipicephalus appendiculatus, and Rhipicephalus sanguineus. They attached to and actively penetrated the cells. Cells that were penetrated by or infected with B. caballi showed rounding off, degeneration, and lysis. In Giemsa-stained spreads prepared after 1 or 2 days in vitro, more kinetes were associated with R. appendiculatus cells when the medium contained A. nitens egg extract (TEE). Ninety percent of the B. caballi kinetes were always found within or in close contact with cells from A. nitens, cultured with or without TEE. The parasites were club shaped, amoeboid, or round. Intracellular Babesia were frequently lying adjacent to the host cell nucleus. In the cultures the parasites were detected by light microscopy for 3 to 5 days; they did not transform into other tick-associated stages and were not seen after 1 week. This is the first report of the cultivation of tick stages of B. caballi in tick cell cultures.     

Identification of NbME MITE families: Potential molecular markers in the microsporidia Nosema bombycis

Six novel families of miniature inverted-repeat transposable elements (MITEs) were characterized in the microsporidia Nosema bombycis and were named NbMEs. The structural characteristics and the distribution of NbME copies in the N. bombycis genome were investigated, and it was found that portions of NbMEs are associated with gene sections. Potential molecular markers for various N. bombycis strains were identified in this study through utilization of the MITE-AFLP technique. Three distinct pathogenic isolates collected from different areas were distinguished, and polymorphisms were detected using the NbME5 marker, thereby establishing this NbME as a potential marker for studying isolate variation in N. bombycis.     

Eimeria acervulina, E. brunetti, and E. maxima: pathogenic effects of single or mixed infections with low doses of oocysts in chickens.

The pathogenic effects of E. acervulina, E. brunetti, and E. maxima were modified when chickens received mixed infections with these species.Six-week-old chickens were inoculated with doses of 20,000 oocysts of E. acervulina, 1250 oocysts of E. brunetti, and 5000 oocysts of E. maxima given as a single or mixed infection.Typical signs of coccidiosis were apparent in chickens infected with a single Eimeria sp. When birds were given infections composed of two species, the weight loss was greater than that due to either given alone but when three species were given, weight loss was slightly less than that due to infection will E. brunetti alone. Oocyst production due to E. acervulina tended to be similar in birds given this species alone or with E. brunetti. Output fell to less than 50% when E. acervulina was administered with E. maxima. Oocyst production due to E. brunetti and E. maxima decreased when these species were inoculated together and when they were administered with E. acervulina. Lesions of E. acervulina and E. brunetti were superimposed on those of E. maxima in birds given mixed infections.Growth retardation was not evident in chickens inoculated with E. acervulina alone, although weight loss increased when this species was administered with either E. brunetti or E. maxima.     

Fatal penetrating skin ulcers in laboratory-reared octopuses

Young Octopus joubini and O. briareus (35 to 60 days old) that were being reared in high-density groups for biomedical studies developed skin ulcers, whereas octopuses reared in individual containers in the same culture system were disease-free. The ulcers first affected the epidermis of the mantle, and then penetrated downward through the dermis and underlying muscle tissue. Four stages of ulceration were observed. Untreated octopuses with ulcers always died, usually within 4 days. Five species of bacteria were isolated from ulcers: Vibrio alginolyticus, V. damsela, Pseudomonas stutzeri, and Aeromonas caviae from O. joubini; and V. parahaemolyticus, V. damsela, and P. stutzeri from O. briareus. Bacteria were found during all stages of the ulceration. Healthy O. joubini were infected experimentally with four species of bacteria, and V. alginolyticus produced skin ulcers within 2 days. The ulceration was treated with nifurpirinol, and complete healing of the skin occurred within 2 months. The ulcers were probably species-specific because O. maya and O. bimaculoides that were reared in the same culture systems were not affected. The cause of the ulceration was probably an increase in contact among crowded octopuses that produced skin abrasions which were invaded by opportunistic bacteria.     

Identification of Frankia Strains by Two-Dimensional Polyacrylamide Gel Electrophoresis

Fifteen Frankia strains from five different plant species were analyzed by two-dimensional polyacryl-amide gel electrophoresis to determine their relatedness by comparing the polypeptide patterns obtained. Three major subgroups (A, C, and D) were found in the Alnus-Comptonia-Myrica cross-inoculation group. An isolate from Purshia tridentata had a unique protein pattern and represents a distinct group of frankiae. Members of group A were isolated from root nodules of Alnus incana subsp. rugosa and Alnus viridis subsp. crispa. Group C organisms were from A. incana subsp. rugosa and Comptonia peregrina nodules, and group D organisms were from A. incana subsp. rugosa, A. viridis subsp. cripsa, and Myrica pensylvanica root nodules. Isolates from each gel group were obtained at several widely separated geographical locations. The results indicate that two-dimensional polyacrylamide gel electrophoresis is useful for identifying Frankia isolates.     

Characterization of novel 2-oxoglutarate dependent dioxygenases converting l-proline to cis-4-hydroxy-l-proline

Hydroxyprolines are valuable chiral building blocks for organic synthesis of pharmaceuticals. Several microorganisms producing l-proline trans-4- and cis-3-hydroxylase were discovered and these enzymes were applied to the industrial production of trans-4- and cis-3-hydroxy-l-proline, respectively. Meanwhile, other hydroxyproline isomers, cis-4- and trans-3-hydroxy-l-proline, were not easily available because the corresponding hydroxylase have not been discovered. Herein we report novel l-proline cis-4-hydroxylases converting free l-proline to cis-4-hydroxy-l-proline. Two genes encoding uncharacterized proteins from Mesorhizobium loti and Sinorhizobium meliloti were cloned and overexpressed in Escherichia coli, respectively. The functions of purified proteins were investigated in detail, and consequently we detected l-proline cis-4-hydroxylase activity in both proteins. Likewise l-proline trans-4-, cis-3-hydroxylase and prolyl hydroxylase, these enzymes belonged to a 2-oxoglutarate dependent dioxygenase family and required a non-heme ferrous ion. Although their reaction mechanisms were similar to other hydroxylases, the amino acid sequence homology was not observed (less than 40%).     

Soil pH Effects on Nematode Populations Associated with Soybeans

''Amsoy'' soybeans were grown for 2 months in nonsterilized Jackson silt loam amended to pH 4.0, 6.0, and 8.0. Nematodes were extracted biweekly from soil and roots. The greatest numbers of Pratylenchus alleni colonized soybean roots at pH 6.0. Hoplolaimus galeatus and members of the Tylenchinae-Psilenchinae survived best at pH 6.0, while numbers o f the Dorylaimoidea were greatest at both pH 6.0 and 8.0. The non-stylet nematodes were recovered in greater numbers from pH 8.0 soil. Potassium, manganese, and phenols were highest in soybean plants grown in pH 4.0 soil, the pH at which there were the fewest nematodes. A thicker suberized outer layer o f root tissue occurred in plants grown at pH 4.0.