Partial 16S rRNA Gene Sequences Research Articles

Is the gut microbiota dysbiotic in patients with classical homocystinuria?

Classical homocystinuria (HCU) is characterized by increased plasma levels of total homocysteine (tHcy) and methionine (Met). Treatment may involve supplementation of B vitamins and essential amino acids, as well as restricted Met intake. Dysbiosis has been described in some inborn errors of metabolism, but has not been investigated in HCU. The aim of this study was to investigate the gut microbiota of HCU patients on treatment. Six unrelated HCU patients (males = 5, median age = 25.5 years) and six age-and-sex-matched healthy controls (males = 5, median age = 24.5 years) had their fecal microbiota characterized through partial 16S rRNA gene sequencing. Fecal pH, a 3-day dietary record, medical history, and current medications were recorded for both groups. All patients were nonresponsive to pyridoxine and were on a Met-restricted diet and presented with high tHcy. Oral supplementation of folate (n = 6) and pyridoxine (n = 5), oral intake of betaine (n = 4), and IM vitamin B12 supplementation (n = 4), were reported only in the HCU group. Patients had decreased daily intake of fat, cholesterol, vitamin D, and selenium compared to controls (p < 0.05). There was no difference in alpha and beta diversity between the groups. HCU patients had overrepresentation of the Eubacterium coprostanoligenes group and underrepresentation of the Alistipes, Family XIII UCG-001, and Parabacteroidetes genera. HCU patients and controls had similar gut microbiota diversity, despite differential abundance of some bacterial genera. Diet, betaine, vitamin B supplementation, and host genetics may contribute to these differences in microbial ecology.

SXT int harboring bacteria as effective indicators to determine high-risk reservoirs of multiple antibiotic resistance in different aquatic environments of western coast of Gujarat, India

Unregulated use of antibiotics in the developing countries has led to the development of Multiple Antibiotic Resistant (MAR) strains. Resistance transferring elements like SXT play crucial role in dissemination of the resistance genes. Persistence of antibiotics and their resistance elements in an aquatic ecosystem depends on the physicochemical properties, and thus may depend on the type of whole water body in general. The study focuses on isolation of bacteria possessing SXT integrase from diverse aquatic bodies like polluted sources, freshwater ponds, groundwater and seawaters of western Gujarat coast, and studying distribution of multidrug resistance in them. Initially, 246 Co-Trimoxazole resistant microbes were isolated from various environmental samples on the basis of morphological distinction. In the PCR analysis, 45 of them were found to be positive for the SXT integrase gene and subsequent analysis confirmed the presence of SXT or SXT-variants. Partial 16S rRNA gene sequencing and diversity analysis on the basis of Shannon’s diversity indices revealed bacterial class-wise diversity trend to be pond water > seawaters > ground water > polluted sources. Antibiogram of these organisms revealed the order of antibiotic resistance indices to be polluted sources > groundwater sources > freshwater ponds > seawater sources. However, the trend of percentage resistance towards antibiotic classes separately in the studied aquatic environments was groundwater sources > polluted sources > freshwater ponds > seawater. Further, the trend for therapeutic efficiency of antibiotic classes was similar in all waters to that of total isolates i.e. Aminoglycosides > Fluoroquinolones > Amphenicols > Cephalosporins > Imidazolidinedione derivatives > Glycopeptide > Quinolones > Macrolides > Penicillins > Sulfa drugs. Positive correlation was observed between prevalence of SXT int positive bacteria and multiple antibiotic resistance incidences; but negative correlation was observed between SXT int positive bacteria and class wise bacterial diversity in four designated type of aquatic environment. Thus, screening for occurrence frequency of SXT integrase gene in the bacterial isolates, may prove as an effective parameter to estimate the multiple antibiotics’ resistance indices in any of the four studies aquatic ecosystems, and thereby determine the antibiotic pollution resulting in the corresponding pressure. The overall quantitative values were logically higher in polluted sources in terms of total resistance indices, and freshwater ponds in terms of diversity. But the groundwater sites may be regarded as the prominent high-risk zones of contamination with multiple antibiotic and/or their resistance traits qualitatively.

Leptospiral Culture without 5’-Fluorouracil Revealed Improved Leptospira Isolation from Febrile Patients in North-Eastern Malaysia

Objectives: Isolation of Leptospira by culture represents a definitive growth and confirmation of the disease, yet it is hampered with its nature of slow growth. With slight modification of culture method, the study aims to isolate and characterize Leptospira spp. from patients with acute febrile illness. Methods: A total of 109 blood samples were collected from patients with acute febrile illness that presented at the Emergency Department of Hospital Universiti Sains Malaysia, Malaysia. Clinical samples were subjected to Leptospira IgM Rapid test, microscopic agglutination test (MAT), isolation by culture method, and direct real-time PCR test. For leptospiral isolation, the samples (whole blood and deposit from spun plasma) were cultured into modified Ellinghausen McCullough Johnson Harris (EMJH) media with and without 5’-fluorouracil (5-FU). In every culture positive sample, partial 16S rRNA gene sequencing was performed for molecular identification of the isolates. Phylogenetic analysis was carried out to determine the genetic relatedness among the isolates. An inhibition of 5-FU study was performed on Leptospira interrogans serovar Canicola with different concentrations to compare the growth detection of the tested Leptospira with or without 5-FU within 7 days of incubation. Results: Leptospirosis was diagnosed in 14.7% of patients with acute febrile illness. Two Leptospira spp. (n = 2/109, 1.85%) were successfully isolated from whole blood and deposit from spun plasma samples. B004 and B208 samples were positive at day 11 and day 7, respectively, in EMJH media without addition of 5-FU. Sample B004 was identified as Leptospira interrogans and B208 as Leptospira weilli. Phylogenetic analysis confirmed that both of them were within pathogenic group and they were not related. The 5-FU inhibition study revealed that additional of 5-FU at final concentration of 200 µg/mL to EMJH media demonstrated an inhibitory effect on the growth of the tested strain Conclusion: Isolation of Leptospira spp. using EMJH media without addition of 5’-fluorouracil resulted in a better outcome. Two pathogenic Leptospira isolates were successfully cultivated from patients with acute febrile illness that were genetically not related.

Photobacterium lucens sp. nov., Isolated from a Cultured Shrimp Penaeus vannamei.

Two bacterial strains were isolated from the hepatopancreas of a cultured shrimp (Penaeus vannamei) in Sinaloa, México. Their partial 16S rRNA gene sequences clustered within those of the genus Photobacterium, showing high similarity to the type strains of Photobacterium angustum and Photobacterium leiognathi, were 87.1% and 97.5%, respectively. Multilocus sequence analysis using eight housekeeping genes (ftsZ, gapA, gyrB, mreB, pyrH, recA, rpoA, topA and 16S rRNA) and phylogenetic analysis with 139 single-copy genes showed that the new strains form an independent branch whole genome sequencing and genomic analyses (average nucleotide identity, average amino acid identity, and in silico DNA-DNA hybridization) produced values well below the thresholds for species delineation with all methods tested. In addition, a phenotypic characterization was performed to support the description and differentiation of the novel strains from related taxa. The results obtained demonstrate that the two strains represent a novel species for which the name Photobacterium lucens sp. nov. is proposed.

Khat Chewing Induces a Floral Shift in Dental Material-Associated Microbiota: A Preliminary Study.

BackgroundThe habit of khat (Catha Edulis Forskal) chewing is widely practiced in the southern regions of Saudi Arabia, Yemen, and East Africa. This social habit has tremendous effects on oral and general health of khat chewers. Khat may affect bacterial species in plaque biofilms on oral rehabilitation materials. This preliminary case-controlled study aimed to assess and compare the effect of khat chewing on bacterial biodiversity between non-khat chewers (NKC) and khat chewers (KC) in oral biofilms on oral rehabilitation materials using polymerase chain reaction (PCR).Material/MethodsFifty participants were organized into 2 equal groups of NKC and KC, each containing 5 subgroups related to filling material type. Some participants had amalgam (A) or composite (C) restorations, while others had feldspathic porcelain (FP), nickel chromium (NC), and zirconia ceramic (ZC) crowns or bridges. Oral biofilm samples were collected from all participants, DNA was extracted, and samples were subjected to PCR. Bacterial species were then identified and counted. PCR products were sequenced to detect similarity. Partial 16S rRNA gene sequences of the current study samples were compared with 16S rRNA gene sequences from GenBank using BLAST on the National Center for Biotechnology Information website.ResultsThe Streptococcus sp. was the most common bacterial species among our participants (40; 80%), followed by Lactobacillus and Veillonella spp., accounting for 12% (6) and 8% (4), respectively. Streptococcus sp. was observed equally among NKC and KC, but Lactobacillus and Veillonella spp. were higher in KC and NKC, respectively. Lactobacillus was associated mainly with prosthetic materials, and Streptococcus was found among all examined dental restorative materials.ConclusionsThis research concluded that khat chewing significantly affects bacterial biodiversity in oral biofilms in the presence of different restorative and prosthetic dental materials.

Diversity and plant growth-promoting functions of diazotrophic/N-scavenging bacteria isolated from the soils and rhizospheres of two species of Solanum

Studies of the interactions between plants and their microbiome have been conducted worldwide in the search for growth-promoting representative strains for use as biological inputs for agriculture, aiming to achieve more sustainable agriculture practices. With a focus on the isolation of plant growth-promoting (PGP) bacteria with ability to alleviate N stress, representative strains that were found at population densities greater than 104 cells g-1 and that could grow in N-free semisolid media were isolated from soils under different management conditions and from the roots of tomato (Solanum lycopersicum) and lulo (Solanum quitoense) plants that were grown in those soils. A total of 101 bacterial strains were obtained, after which they were phylogenetically categorized and characterized for their basic PGP mechanisms. All strains belonged to the Proteobacteria phylum in the classes Alphaproteobacteria (61% of isolates), Betaproteobacteria (19% of isolates) and Gammaproteobacteria (20% of isolates), with distribution encompassing nine genera, with the predominant genus being Rhizobium (58.4% of isolates). Strains isolated from conventional horticulture (CH) soil composed three bacterial genera, suggesting a lower diversity for the diazotrophs/N scavenger bacterial community than that observed for soils under organic management (ORG) or secondary forest coverture (SF). Conversely, diazotrophs/N scavenger strains from tomato plants grown in CH soil comprised a higher number of bacterial genera than did strains isolated from tomato plants grown in ORG or SF soils. Furthermore, strains isolated from tomato were phylogenetically more diverse than those from lulo. BOX-PCR fingerprinting of all strains revealed a high genetic diversity for several clonal representatives (four Rhizobium species and one Pseudomonas species). Considering the potential PGP mechanisms, 49 strains (48.5% of the total) produced IAA (2.96-193.97 μg IAA mg protein-1), 72 strains (71.3%) solubilized FePO4 (0.40-56.00 mg l-1), 44 strains (43.5%) solubilized AlPO4 (0.62-17.05 mg l-1), and 44 strains produced siderophores (1.06-3.23). Further, 91 isolates (90.1% of total) showed at least one PGP trait, and 68 isolates (67.3%) showed multiple PGP traits. Greenhouse trials using the bacterial collection to inoculate tomato or lulo plants revealed increases in plant biomass (roots, shoots or both plant tissues) elicited by 65 strains (54.5% of the bacterial collection), of which 36 were obtained from the tomato rhizosphere, 15 were obtained from the lulo rhizosphere, and 14 originated from samples of soil that lacked plants. In addition, 18 strains showed positive inoculation effects on both Solanum species, of which 12 were classified as Rhizobium spp. by partial 16S rRNA gene sequencing. Overall, the strategy adopted allowed us to identify the variability in the composition of culturable diazotroph/N-scavenger representatives from soils under different management conditions by using two Solanum species as trap plants. The present results suggest the ability of tomato and lulo plants to enrich their belowground microbiomes with rhizobia representatives and the potential of selected rhizobial strains to promote the growth of Solanum crops under limiting N supply.

Endophytic bacteria of Anisophyllea disticha (Raja Berangkat) from tropical lake environment in Malaysia

Lake Kenyir is the largest artificial lake in South East Asia and serves as habitats for diverse flora and fauna. Moreover, Lake Kenyir is a rich resource of medicinal plants which has been used by local community surrounding the lake. Endophytic bacteria are able to colonize plants without causing harm and they can be highly beneficial towards their plant hosts. The aim of this study is to isolate and identify endophytic bacteria from the leaves and roots of a medicinal plant, Anisophyllea disticha (Raja Berangkat), and to evaluate their antimicrobial capabilities. Twenty isolates each from the leaves and roots were randomly selected for molecular identification using 16S rRNA gene. Partial 16S rRNA gene sequences revealed that genus Bacillus dominated the endophytic bacteria from the leaves with two isolates belonging to Paenibacillus and Aeribacillus. Five genera namely, Bacillus, Staphylococcus, Lysinibacillus, Paenibacillus, and Mycobacterium were successfully obtained from the roots of A. disticha. Nine endophytic bacteria—five Bacillus species (R3, R11, R13, R26, R37), Lysinibacillus (R9), two Staphylococcus species (R6, R27) and Mycobacterium species (R33) from roots were found to show weak antibacterial and/or antifungal activities. Findings from this study provide an insight to endophytic bacteria inhabiting A. disticha and the role they may play in their plant host.

Distribution and recombination of Wolbachia endosymbionts in Korean coleopteran insects

BackgroundWolbachia are among the most prevalent endosymbiotic bacteria and induce reproductive anomalies in various invertebrate taxa. The bacterium has huge impacts on host reproductive biology, immunity, evolution, and molecular machinery. However, broad-scale surveys of Wolbachia infections at the order scale, including the order Coleoptera, are limited. In this study, we investigated the Wolbachia infection frequency in 201 Coleopteran insects collected in Korea.ResultsA total of 26 species (12.8%) belonging to 11 families harbored Wolbachia. The phylogenetic trees of based on partial 16S rRNA gene sequences and partial Wolbachia surface protein (wsp) gene sequences were largely incongruent to that of their hosts. This result confirms that Wolbachia evolved independently from their hosts,ConclusionPhylogenetic trees suggest that complex horizontal gene transfer and recombination events occurred within and between divergent Wolbachia subgroups.

Screening of lactic starter from Tunisian fermented vegetables and application for the improvement of caper (Capparis spinosa) fermentation through an experimental factorial design

PurposeThis study aims at designing a lactic starter for caper fermentation isolated from Tunisian fermented vegetables to improve the process and produce consistent and high-quality product.MethodsIn this study, the lactic starter was isolated by exploring the lactic acid bacteria (LAB) of Tunisian artisanal fermented vegetables. Identification was carried out by partial 16S rRNA gene sequencing. Screening was based on salt tolerance and antagonistic activities against Escherichia coli ATCC 10536 and Enterococcus faecalis ATCC 10541. Caper fermentation was optimized through a full factorial experimental design (23), by exploring three factors: starter inoculum size, NaCl concentration, and acetate content. Differences in pH values, Total aerobic mesophilic bacteria and LAB counts between the beginning and end of fermentation are selected as responses and corresponding regression coefficients were calculated.ResultsThe lactic microbiota is mainly represented by Lactobacillus plantarum group. Based on salt tolerance and antimicrobial activity, the strain Lactobacillus plantarum F3 was selected as starter for caper fermentation. The effect of NaCl concentration, acetate content, and inoculum size on acidity, total aerobic mesophilic bacteria count, and LAB count after 1 week and 1 month of caper fermentation was studied. Depending on the fermentation time, either 1 week or 1 month, the initial conditions should comprise 0% acetate, 108 CFU/mL inoculum, and 5% NaCl for 1 week against 5% acetate, 107 CFU/mL inoculum, and 10% NaCl for 1 month lasting caper fermentation. A protocol for caper fermentation was set up ensuring hygienic quality and LAB viability.ConclusionLb. plantarum F3 was selected as lactic starter for caper fermentation, and initial fermentation conditions were optimized through a full factorial design. This work has shown loss in LAB viability after 1 week of fermentation. Based on results obtained, an optimized fermentation protocol was set up. This protocol ensures LAB survival and high hygienic quality of the product.

Mangrove soil as a source for novel xylanase and amylase as determined by cultivation-dependent and cultivation-independent methods.

Xylanase and α-amylase enzymes participate in the degradation of organic matter, acting in hemicellulose and starch mineralization, respectively, and are in high demand for industrial use. Mangroves represent a promising source for bioprospecting enzymes due to their unique characteristics, such as fluctuations in oxic/anoxic conditions and salinity. In this context, the present work aimed to bioprospect xylanases from mangrove soil using cultivation-dependent and cultivation-independent methods. Through screening from a metagenomic library, three potentially xylanolytic clones were obtained and sequenced, and reads were assembled into contigs and annotated. The contig MgrBr135 was affiliated with the Planctomycetaceae family and was one of 30 ORFs selected for subcloning that demonstrated only amylase activity. Through the cultivation method, 38 bacterial isolates with xylanolytic activity were isolated. Isolate 11 showed an enzymatic index of 10.9 using the plate assay method. Isolate 39 achieved an enzyme activity of 0.43 U/mL using the colorimetric method with 3,5-dinitrosalicylic acid. Isolate 39 produced xylanase on culture medium with salinity ranging from 1.25 to 5%. Partial 16S rRNA gene sequencing identified isolates in the Bacillus and Paenibacillus genera. The results of this study highlight the importance of mangroves as an enzyme source and show that bacterial groups can be used for starch and hemicellulose degradation.

&lt;p class="Body"&gt;&lt;strong&gt;Detection of &lt;em&gt;Theileria luwenshuni&lt;/em&gt; (Piroplasmida: Theileriidae) from ticks infesting goats in peninsular Malaysia&lt;/strong&gt;&lt;/p&gt;

Piroplasmic parasites such as Theileria and Babesia commonly infect important livestock animals, leading to economic losses in the livestock industry. These parasites are primarily transmitted by ticks found infesting livestock animals. In Malaysia, livestock diseases caused by tick-borne pathogens are still under-studied, even though tick infestations are common in many livestock farms. Here, we molecularly detect a Theileria sp. from Haemaphysalis bispinosa ticks infesting goats in a privately-owned farm in Perak, peninsular Malaysia. PCR was performed on the DNA extracted from ticks to determine the presence of piroplasms. Three ticks were found PCR-positive for piroplasms. Sequence analyses of partial 18S rRNA gene sequences revealed the presence of a Theileria species at 100% identity to Theileria luwenshuni. Our findings suggest the presence of T. luwenshuni, a highly pathogenic Theileria sp. to sheep and goats, in the small ruminant industry in Malaysia.

Isolation, identification and potential biological control of some rhizobacteria against Meloidogyne incognita

The plant-parasitic nematodes seriously effect on the growth of many crops and are responsible for agricultural losses worldwide. The losses range from 8 to 20% on major crops around the world. Root-knot nematode Meloidogyne spp. infect and damage a wide range of important crops particularly vegetables in tropical and subtropical countries. The main way for controlling of the plant parasitic nematodes is the use of chemical nematicides. Although the nematicides are effective quickly, they are usually expensive and not available and also cause a lot of risk to humans and inflict injury to the environment. Due to the environmental hazards associated with their application, identifying alternatives for nematode control and developing effective and safe application techniques is both urgent strategies for alleviating the nematode induced damage. The biological controls are alternatives eco-friendly agricultural systems and safer for environments and humans and cheaper than chemical control This study was conducted to isolate and estimate of the potential some rhizobacteria from roots in agriculture soil against root-knot nematode, Meloidogyne incognita J2 under in vitro condition. Nine rhizobacteria were isolated from soil (R1 to R9). The best mortality was recorded by isolate no. R6 (77.55%) followed by isolated no. R2 (75.59%) followed by isolate no. R7 (71.43%) as compared with 0% in case of control (water only) after 72h exposure periods. R6, R2, and R7 were identified as Lysinibacillus sphaericus, Bacillus pumilus and Pseudomonas flourescens, respectively based on the analysis of the 16S rRNA gene sequence. The partial 16S rRNA gene sequence of these bacterial isolates were deposited in GenBank under accession number MF000302, MF000303 and MF000304 for the previous mentioned bacterial species. The main goals of this study were to isolate some native rhizobacteria and evaluate their efficiency as eco-friendly control alternatives for controlling root- knot nematode M. incognita under laboratory conditions.

Variation in the microbial community contributes to the improvement of the main active compounds of Magnolia officinalis Rehd. et Wils in the process of sweating

BackgroundMagnolia officinalis Rehd. et Wils, commonly called Houpo, has been used for thousands of years in China as a traditional herbal medicine. The primary processing of Houpo requires sweating treatment, which is a special drying process and is considered to be an essential embodiment of high quality and genuine medicinal materials. The sweating of Houpo leads to peculiar changes in the microbial community structure and the content of main active substances (magnolol, honokiol, syringin and magnoflorine). Variation in the microbial community was considered the cause of the change in content of active substances of Houpo, although the microbial taxa responsible for the improvement of content remain unidentified.MethodsIn this study, we used MiSeq high-throughput sequencing methods for partial bacterial 16S rRNA and 18S rRNA gene sequences to compare the bacterial and fungal community structures at different timepoints in the process of sweating. The content of the main active substances (magnolol, honokiol, syringin and magnoflorine) were determined by high-performance liquid chromatography analysis to evaluate the effects of sweating. UPLC-Q-Extractive Orbitrap mass spectrometry (UPLC-QE Orbitrap MS) was used to detection of differential metabolites of unsweated Houpo before and after co-culture with core bacterial solutions.ResultsIn this study, the total contents of magnolol (MG) and honokiol (HK) were significantly increased at 4 dp (dp for day PM sample), up to 3.75%, and the contents of syringin (SG) and magnoflorine (MF) were as high as 0.12% and 0.06%, respectively. Bacterial abundance and diversity were higher in the early stage (0 day–2 da; da for day AM sample) than in the later stage (4–5 dp), while fungal abundance was more obvious in the later stage than in the early stage. Positive correlation coefficients revealed that the relative abundance of Enterobacter (P < 0.05), Klebsiella (P < 0.05), Weissella (P < 0.05), Bacillus (P < 0.05) and Candida (P < 0.05) would be conducive to improving the quality of Houpo. Negative correlation coefficients revealed that the relative abundance of Actinomycetospora, Singulisphaera, Mucilaginibacter, Deinococcus, Gemmatirosa, Methylobacterium, Sphingomonas, Hymenobacter, Halomonas and Capnobotryella could be a potential antagonist for the decrease in the quality of Houpo. After co-culture of single core strain and unsweated Houpo, there was no significant difference in the four main active components, but there were other metabolites with significant difference.ConclusionsOur findings reveal that sweating increased the content of the main active compounds, promoted the relative abundance of potentially beneficial microbes, decreased the abundance of potentially harmful microbes, the core functional genera group together, forming a core microbiome, these genera are dominant across the different stages of the sweating process and contribute to the quality development of the characteristics of Houpo. Meanwhile, this study presented a clear scope for potential beneficial microbes that improve the quality of Houpo.

Molecular Detection and Phylogenetic Analysis of Anaplasma Spp. in Cattle in Al-Qadisiyah Province of Iraq

Abstract Anaplasma spp. are widely spread rickettsial bacteria transmitted by ticks and placing high impacts on veterinary and public health. A limited number of studies have been carried out on Anaplasmosis in the central part of Iraq. This study was conducted to determine the presence of Anaplasma spp. in cattle in Al-Qadisiyah province, Iraq. A total of 400 blood specimens were collected from cattle suffering from heavy tick infestation. Cattle were blood-sampled from four hyper-endemic areas with ticks. Blood samples were screened using microscopic and polymerase chain reaction (PCR) methods. Diff-quick stained blood smears revealed Anaplasma-like inclusion bodies in 254 (63.5%) samples. According to the 16S rRNA-gene-based PCR analysis, Anaplasma spp. was detected in 124 of the 400 (31%) samples, divided as 96/254 (37.8%) among the microscopical positive samples and 28/146 (19.17%) among the microscopical negative samples. Phylogenetic analysis based on the partial 16S rRNA gene sequencing of ten-PCR positive samples were 99–97% identical to sequences deposited in the GenBank, revealing presence of A. phagocytophilum, A. marginale and unnamed Anaplasma spp. in 40%, 20%, and 40% samples, respectively. Relationships among Anaplasma spp. infections and cattle breed, age, and sex were analyzed. Calves less than one year old showed significantly higher rates (p&lt;0.005) than those from other age groups, whereas sex and breed demonstrated no significant differences (p˃0.001). This study shows that a variety of Anaplasma spp., were endemic in central part of Iraq and is still a hidden problem in cattle in the hyperendemic areas of tick, which requires serious control strategies.

Plant variety and soil type influence Escherichia coli O104:H4 strain C227/11ϕcu adherence to and internalization into the roots of lettuce plants

Human disease outbreaks caused by pathogenic Escherichia coli are increasingly associated with the consumption of contaminated fresh produce. Internalization of enteroaggregative/enterohemorrhagic E. coli (EAEC/EHEC) strains into plant tissues may present a serious threat to public health. In the current study, the ability of the fluorescing Shiga toxin-negative E. coli O104:H4 strain C227/11ϕcu/pKEC2 to adhere to and to internalize into the roots of Lactuca sativa and Valerianella locusta grown in diluvial sand (DS) and alluvial loam (AL) was investigated. In parallel, the soil microbiota was analyzed by partial 16S rRNA gene sequencing. The experiments were performed in a safety level 3 greenhouse to simulate agricultural practice. The adherence of C227/11ϕcu/pKEC2 to the roots of both plant varieties was increased by at least a factor three after incubation in DS compared to AL. Compared to V. locusta, internalization into the roots of L. sativa was increased 12-fold in DS and 108-fold in AL. This demonstrates that the plant variety had an impact on the internalization ability, whereas for a given plant variety the soil type also affected bacterial internalization. In addition, microbiota analysis detected the inoculated strain and showed large differences in the bacterial composition between the soil types.

Plant growth promoting rhizobacteria and their potential as bioinoculants on Pennisetum clandestinum (Poaceae)

In this study, we aimed at examining the potential to stimulate growth in Pennisetum clandestinum using four isolated bacterial strains from soils obtained from a Colombian tropical silvopastoral system. We previously identified genetically the strains and characterized two plant growth promotion activities. We found that the four bacterial strains were phylogenetically associated with Klebsiella sp. (strains 28P and 35P), Beijerinka sp. (37L) and Achromobacter xylosoxidans (E37), based on partial 16S rRNA gene sequencing. Moreover, the in vitro biochemical assays demonstrated that the strains exhibited some plant growth promotion mechanisms such as 1-aminocyclopropane-1-carboxylic acid deaminase activity and indole compound synthesis. Notably, bacterial inoculation under greenhouse conditions showed a positive influence on P. clandestinum growth. We found a significant (p &lt; 0.05) effect on root and shoot length, and shoot dry weight. Shoot length increased by 52% and 30% with 37L and 35P, respectively, compared to those without inoculation treatment. Similarly, the use of 37L and 28P raised shoot dry weight values by 170% and 131%, respectively. In root development, inoculation with strains 37L and E37 increased root length by 134% and 100%, respectively. Beijerinckia sp. 37L was the most effective of the four strains at increasing P. clandestinum biomass and length.

THE DIVERSITY OF CULTIVABLE HYDROCARBON-DEGRADING BACTERIA ISOLATED FROM CRUDE OIL CONTAMINATED SOIL AND SLUDGE FROM ARZEW REFINERY IN ALGERIA

The use of autochtonious bacterial strains is a valuable bioremediation strategy for cleaning the environment from hydrocarbon pollutants. The isolation, selection and identification of hydrocarbon-degrading bacteria is therefore crucial for obtaining the most promising strains for decontaminate a specific site. In this study, two different media, a minimal medium supplemented with petroleum and with oil refinery sludge as sole carbon source, were used for the isolation of native hydrocarbon-degrading bacterial strains from crude oil contaminated soils and oil refinery sludges which allowed isolation of fifty-eight strains.The evalution of diversity of twenty-two bacterials isolates reveled a dominance of the phylum Proteobacteria (20/22 strains), with a unique class of Alphaproteobacteria, the two remaining strains belong to the phylum Actinobacteria. Partial 16S rRNA gene sequencing performed on isolates showed high level of identity with known sequences. Strains were affiliated to Sinorhizobium, Promicromonospora, Novosphingobium, Georgenia, Ancylobacter, Roseomonas, Hansschlegelia and Tistrella genera. Research for the genes that encoding for degradation enzymes in isolated genera genome data deposited in Genbank reveled the presence of degradation gene in three species Sinorhizobium meliloti, Novosphingobium panipatenseand Tistrella mobilis.

First Report of Leaf Blight of Rice Caused by Pantoea ananatis and Pantoea dispersa in Malaysia

HomePlant DiseaseVol. 103, No. 7First Report of Leaf Blight of Rice Caused by Pantoea ananatis and Pantoea dispersa in Malaysia PreviousNext DISEASE NOTESFirst Report of Leaf Blight of Rice Caused by Pantoea ananatis and Pantoea dispersa in MalaysiaW. K. Toh, P. C. Loh, and H. L. WongW. K. TohFaculty of Science, Universiti Tunku Abdul Rahman, 31900 Kampar, MalaysiaSearch for more papers by this author, P. C. LohFaculty of Science, Universiti Tunku Abdul Rahman, 31900 Kampar, MalaysiaSearch for more papers by this author, and H. L. Wong†Corresponding author: H. L. Wong; E-mail Address: hlwong@utar.edu.myhttp://orcid.org/0000-0001-8903-1812Faculty of Science, Universiti Tunku Abdul Rahman, 31900 Kampar, MalaysiaSearch for more papers by this authorAffiliationsAuthors and Affiliations W. K. Toh P. C. Loh H. L. Wong † Faculty of Science, Universiti Tunku Abdul Rahman, 31900 Kampar, Malaysia Published Online:16 May 2019https://doi.org/10.1094/PDIS-12-18-2299-PDNAboutSectionsSupplemental ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinked InRedditEmailWechat In September to December 2016, the most severe leaf blight disease outbreak in the last 30 years occurred in the paddy field of Sekinchan, Selangor, Malaysia, causing 50 to 70% loss of the crop yield. During this outbreak, the newly introduced rice cultivar MR 284 suffered the most damage. Infected rice leaf blades showed brownish stripes, which subsequently turned pale and dry. Symptomatic leaves from the MR 284 plants were cut into 1-cm2 pieces, surface sterilized, and plated on peptone sucrose agar (Poulin et al. 2014). Yellowish bacterial colonies that were obtained after incubation at 28°C for 2 days were purified by streaking on nutrient agar. Identification of six randomly chosen bacterial isolates was performed using the BioMérieux Vitek 2 microbial identification system, which classified the isolates as Pantoea spp. with 93 to 96% identity probabilities. Subsequently, 16S rRNA gene amplification was performed using the universal primers 27F and 1492R. BLASTn analysis of the partial 16S rRNA gene sequences from the six isolates revealed that the sequences (1,421 bp) from two isolates, designated as PA strains, were identical to those of Pantoea ananatis (GenBank accession no. CP028033.1), and the sequences (1,435 bp) of four isolates, designated as PC strains, were identical to those of Pantoea dispersa (GenBank accession no. AB273743.1). The partial 16S rRNA gene sequences of PA and PC strains were deposited at GenBank under accession numbers MK578186 and MK263020, respectively. Pathogenicity assays of PA and PC strains were conducted on 60-day-old rice plants of cultivar MR 284 using the clipping method (Kauffman et al. 1973). The bacterial strains were cultured overnight on nutrient broth, and the inocula were adjusted to 108 cells/ml. Three fully extended leaves from the second and third tillers of each plant were inoculated by cutting at 3 to 4 cm from the leaf tip. Control plants were mock inoculated with sterile saline solution. The experiment was performed in duplicate. From 3 to 14 days after bacterial inoculation, the inoculated leaves showed progressive necrosis with color change from pale green, brown, and pale, whereas the control plants remained symptomless. Bacterial colonies were reisolated from inoculated leaves, producing colonies that were morphologically identical to those of PA and PC strains. The identities of these bacterial isolates were further verified to be the inocula PA and PC strains by 16S rRNA gene sequencing, thus fulfilling the Koch’s postulates. Previous reports had established Pantoea spp. as rice pathogens (Kini et al. 2017; Lee et al. 2010; Mondal et al. 2011). To our knowledge, this is the first report of rice leaf blight caused by P. ananatis and P. dispersa in Malaysia. The isolation of these pathogens allows newly developed cultivars to be more adequately tested before these cultivars are introduced to the farmers for mass cultivation.The author(s) declare no conflict of interest.

Report of Bursaphelenchus crenati Rühm, 1956 (Nematoda: Parasitaphelenchinae) from Belarus and Russia with a diagnostic key and phylogeny of the Sexdentati group

AbstractThe endoparasitic nematode, Bursaphelenchus crenati, in beetle tunnels of Hylesinus crenatus from the ash Fraxinus excelsior L. with signs of ash dieback, is recorded for the first time in Central Russia and Belarus. Third‐stage dauer juveniles were extracted from the bark beetle, H. crenatus, and cultured on Botrytis cinerea. Morphological, morphometric and molecular analyses of these populations are presented here. Some morphological differences of dauers and adults were found between Belarusian and Russian populations, which may be attributed to seasonal variations. For description of new Bursaphelenchus species, it is recommended to indicate not only the number of lateral incisures but also the number of bands. Recommendations are made to classify the male caudal papillae of Bursaphelenchus into two groups: papilliform and gland papillae with separate numerals. Emended diagnosis, tabular diagnostic polytomous key for the species of Sexdentati group with lists of their vectors and plants, is given. Phylogenetic relationships between some species of the Sexdentati group are also presented based on the analysis of partial 18S, ITS and 28S rRNA gene sequences.

Studies on Antibacterial Activity and Diversity of Cultivable Actinobacteria Isolated from Mangrove Soil in Futian and Maoweihai of China.

Mangrove is a rich and underexploited ecosystem with great microbial diversity for discovery of novel and chemically diverse antimicrobial compounds. The goal of the study was to explore the pharmaceutical actinobacterial resources from mangrove soil and gain insight into the diversity and novelty of cultivable actinobacteria. Consequently, 10 mangrove soil samples were collected from Futian and Maoweihai of China, and the culture-dependent method was employed to obtain actinobacteria. A total of 539 cultivable actinobacteria were isolated and distributed in 39 genera affiliated to 18 families of 8 orders by comparison analysis of partial 16S rRNA gene sequences. The dominant genus was Streptomyces (16.0 %), followed by Microbacterium (14.5 %), Agromyces (14.3 %), and Rhodococcus (11.9 %). Other 35 rare actinobacterial genera accounted for minor proportions. Notably, 11 strains showed relatively low 16S rRNA gene sequence similarities (< 98.65 %) with validly described species. Based on genotypic analyses and phenotypic characteristics, 115 out of the 539 actinobacterial strains were chosen as representative strains to test their antibacterial activities against “ESKAPE” bacteria by agar well diffusion method and antibacterial mechanism by the double fluorescent protein reporter system. Fifty-four strains in 23 genera, including 2 potential new species, displayed antagonistic activity in antibacterial assay. Meanwhile, 5 strains in 3 genera exhibited inhibitory activity on protein biosynthesis due to ribosome stalling. These results demonstrate that cultivable actinobacteria from mangrove soil are potentially rich sources for discovery of new antibacterial metabolites and new actinobacterial taxa.