16S rDNA Sequence Analysis Research Articles

Contribution of tick-borne diseases to mortality in juvenile free-living cervids

Reports on tick-borne infections in free-living juvenile animals and their impact on survival of cervids in nature are lacking. The aim of the study was to detect and identify the Babesia and Anaplasma phagocytophilum species/ecotypes that may have contributed to the death of juvenile animals from a wildlife rescue centre in spring 2020.PCR amplification and sequencing of two genetic markers (18S rDNA and cox1 for Babesia, 16S rDNA and groEL for A. phagocytophilum) were used for screening eleven samples derived from juvenile animals which died in a rescue centre (seven roe deer Capreolus capreolus, one elk Alces alces, one red squirrel Sciurus vulgaris, one European beaver Castor fiber, one red fox Vulpes vulpes). Phylogenetic analysis of full-length 18S rDNA sequence was performed to enable differentiation between two closely-related species infecting wild ungulates, Babesia capreoli and Babesia divergens (zoonotic).The occurrence of the typical SNPs of B. capreoli at two discriminating positions in the 18S rRNA gene allowed identification of B. capreoli infection in a roe deer calf. In two calves, Anaplasma phagocytophilum ecotype 2 was identified, including the same calf co-infection. No Babesia DNA was amplified in an elk calf treated for babesiosis. Splenomegaly was recorded in roe deer calves with A. phagocytophilum and A. phagocytophilum + B. capreoli infection.The study revealed that tick-borne infections are common among juvenile, free-living cervids and may contribute to their mortality.

What Babesia microti Is Now.

Parasites from diverse hosts morphologically identified as Babesia microti have previously been shown to belong to a paraphyletic species complex. With a growing number of reports of B. microti-like parasites from across the world, this paper seeks to report on the current knowledge of the diversity of this species complex. Phylogenetic analysis of 18S rDNA sequences obtained from GenBank shows that the diversity of the B. microti species complex has markedly increased and now encompasses at least five distinct clades. This cryptic diversity calls into question much of our current knowledge of the life cycle of these parasites, as many biological studies were conducted before DNA sequencing technology was available. In many cases, it is uncertain which B. microti-like parasite was studied because parasites from different clades may occur sympatrically and even share the same host. Progress can only be made if future studies are conducted with careful attention to parasite identification and PCR primer specificity.

First Molecular Characterisation of Blastocystis from Experimental Rats in Turkey and Comparison of the Frequencies Between Obese and Non-obese Groups

Blastocystis is a zoonotic protozoan that infects a wide range of animals, including humans and rodents. This study aimed to determine the frequency and subtype distribution of Blastocystis in laboratory rats at a laboratory animal facility in Turkey. This study included 54 male Sprague-Dawley rats from Aydın Adnan Menderes University Laboratory Animal Center. Among these rats, 30 were fed with high-fat diet (obese group) and the remaining 24 received standard chow (non-obese group). Blastocystis positivity was determined with amplification of small subunit 18S rRNA gene following their nucleic acid extraction from faecal samples. Subtypes were detected by submitting the partial 18S rRNA gene sequences to the database (pubmlst. Blastocystis infection was detected in 33 (61.1%) of 54 laboratory rats. The frequency of Blastocystis was significantly different between obese and non-obese rats (p<0.05), with 43.3% and 83.3%, respectively. When referred to the database, exact matches were identified with Blastocystis subtype 4 (ST4) for all isolates. In the phylogenetic analysis of the partial 18S rDNA sequence, the sequence was closely clustered with reference ST4 subtypes from other countries, including China, Japan, United Kingdom and Czech Republic. This study revealed the high rate of Blastocystis colonisation in laboratory rats, posing a risk for human transmission. The comparison of obese and non-obese groups supported the idea that Blastocystis might be an indicator of healthy gut flora. The detection of ST4 in all rats agreed with previous reports of the predominance of this subtype in rodents.

Isolation, characterization and identification of novel broad spectrum bacterial antagonist(s) to control Fusarium wilt of eggplant

Soil bacteria are now-a-days familiar for plant disease prevention in comparison to chemical control measures in an eco-friendly way. Antagonistic soil bacteria have created tremendous interests among scientists to be an alternative means of plant disease control. The aim of our study was to isolate soil bacteria with a multifaceted role, both in the aspects of plant nutrition and disease prevention. To achieve our goal, rhizobacteria from crop plants, cultivated in different regions of West Bengal were isolated and from a total of 250 isolates, only eight were primarily selected based on their initial growth suppression capabilities against the phytopathogen Fusarium oxysporum. Finally, the two bacterial isolates, NC1 and NC2, showed in vitro broad-spectrum antifungal activity against seven potent phytopathogenic fungi, i.e., Fusarium oxysporum, Alternaria alternata, Colletotrichum capsici, Macrophomina phaseolina, Sclerotium hydrophilum, Rhizoctonia solani and Penicillium digitatum, where they suppressed mycelial growth up to 69% over untreated control. The two potent antagonists were screened in vitro for their plant growth promoting (PGP) and antifungal properties where they showed their efficacy to produce many lytic enzymes, indole acetic acid (IAA) and ammonia. The isolates were detected as Gram-positive, endospore, and capsule forming rod-shaped bacteria. On the basis of morphological, physiological, biochemical and BLAST analysis of 16s rDNA sequences, NC1 and NC2 were identified as Bacillus amyloliquefaciens KY568716 and Bacillus velezensis KY568715. When applied on eggplant, both the isolates suppressed Fusarium wilt symptoms and enhanced plant growth significantly. The effects were observed up to 30 days after pathogen inoculation and both the isolates were effective in reducing Fusarium wilt disease symptoms by 57.23% and 64.19% respectively, as compared to pathogen treated plants. They also showed significant enhancement of all plant growth parameters such as root and shoot length, biomass and chlorophyll content. Thus, the two isolates can be used to develop an effective dual function bio-control agents to control plant diseases as well as promoting plant growth.

Iron-reducing - and -proteobacteria isolated from laboratory-scaled heterotrophic feammox bioreactor

Ammonium removal from wastewater is a crucial step in wastewater treatment. Presently employed technologies based on nitrification/denitrification and partial nitritation/anammox principles require oxygen for the nitrification step, and are therefore still not yet fully satisfied with the application practice. In recent years, biological ammonium oxidation coupled with ferric iron reduction (feammox) has been proposed to be responsible for the nitrogen loss in different ecological habitats. Related to the wastewater aspect, the feammox principle has been discussed as an alternative approach for ammonium removal without dependency on oxygen. From a laboratory-scaled feammox bioreactor operated under neutral pH, two bacterial strains FN7 and FN9 were isolated by using the anaerobic Hungate technique. Comparative analyses of 16S rDNA sequences showed that these strains were most closely related to the b-proteobacterium Aciclyphilus denitrificans and the g-proteobacterium Pseudomonas stutzeri, respectively. Although being phylogenetically apart, strains FN7 and FN9 shared several common physiological characteristics that are considered meaningful for the feammox process, i.e. (i) heteroptrophic ammonium oxidation, (ii) denitrification, and (iii) ferric iron reduction. These isolates are proposed to play certain roles in the studied feammox system, contributing to the ammonium removal under heterotrophic feammox condition. The 16S rDNA sequences of strains FN7 and FN9 were available in GenBank under the accession numbers LC474369 and MT568614, respectively.

First report of soft rot of cabbage caused by Pectobacterium wasabiae in Japan.

Cabbage (Brassica oleracea var. capitata) is one of the important vegetables in Japan. In the summer of 2019, some cabbages with soft rot were found in commercial fields in Hokkaido, the northern island in Japan. All diseased plants showed grey to brown discoloration and expanding water-soaked lesions on leaves. We obtained two independent strains (NACAB191 and NACAB192) from diseased leaves. DNA from these strains yielded an expected single size amplicon with the primer set of PhF/PhR for P. wasabiae (De Boer et al. 2012) by PCR, but did not yield the expected amplicon with the primer set of BR1f/L1r for P. carotovorum subsp. brasiliense (Duarte et al. 2004) and Eca1f/Eca2r for P. atrosepticum (De Boer et al., 1995) by PCR. These two strains grew at 37°C, and their ability to utilize raffinose and lactose. These bacterial strains were gram-negative and rod-shaped. The bacterium was positive for O-nitrophenyl-beta-D-galactopyranoside, N-acetylglucosaminyl transferase, gelatin liquefaction, and acid production from D-galactose, lactose, melibiose, raffinose, citrate, and trehalose. The bacterium was negative for indole production and acid production from maltose, α-methyl-D-glucoside, sorbitol, D-arabitol, inositol, inulin, and melezitose. All strains exhibited pectolytic activity on potato slices. The sequence analysis of 16S rDNA (LC597897 and LC597898) showed more than 98% identities to P. wasabiae strain (e.g. HAFL01 in Switzerland) by BLAST analysis. In addition, Multi-locus sequence analysis (Ma et al. 2007) was performed by MEGA10 (Kumer et al. 2018) using concatenated DNA sequences of seven housekeeping genes (aconitate hydratase(acnA, LC597923 and LC597924), glyceraldehyde-3-phosphate dehydrogenase A(gapA, LC597970 and LC597971), isocitrate dehydrogenase (icdA, LC597996 and LC597997), malate dehydrogenase(mdh, LC598022 and LC598023), mannitol-1-phosphate dehydrogenase (mtlD, LC598048 and LC598049), glucose-6-phosphate isomerase (pgi, LC598074 and LC598075) and gamma-glutamyl phospate reductase (proA, LC598079 and LC598080)), and all clustered NACAB191 and NACAB192 into a clade containing other confirmed strains of P. wasabiae. As a result, these two strains shared high identity with each other (>98%, E-Values showed 0). The clade containing these two strains was consistently placed in a larger clade with the other P. wasabiae and 100% bootstrap support for its separation from other Pectobacterium species available in GenBank when the consensus tree constructed using Maximum Likelihood method. Pathogenicity of these strains against cabbage (cv. 'Rakuen') was confirmed by the field experiments with five weeks growth plants sprayed with bacterial suspension (1×107cfu/ml). Thirty cabbages per strain were used in this study, 12 plants treated the suspension of NACAB191 and 16 plants treated the suspension of NACAB192 which died with the same soft rot symptoms about four weeks after inoculation. Whereas water-inoculated plants remained symptomless. Strains re-isolated from the artificially diseased stems were confirmed as P. wasabiae using the methods as biochemical characterization and multiple genetic analyses. Based on the disease symptoms, the cultural, molecular, and pathological features of the strains, we conclude that the soft rot symptoms of cabbage in Hokkaido in 2019 were caused by P. wasabiae. To our knowledge, this is the first report of P. wasabiae as the soft rot disease agent of cabbage in Japan.

First report of soft rot of onion caused by Pectobacterium wasabiae in Japan.

Onion (Allium cepa L.) is one of the important vegetables in Japan. In the summer of 2019, onions with soft rot were found in commercial fields in Hokkaido, the northern island in Japan. Diseased onion showed chlorosis, maceration of leaves, and rotted bulbs. We sampled some diseased onions and isolated three independent isolations (NAONI191, NAONI192 and NAONI193) from infected bulbs on LB medium. These strains were identified as Pectobacterium wasabiae based on their inability to grow at 37°C, and their ability to utilize raffinose and lactose. These bacterial strains were gram-negative, rod-shaped, N-acetylglucosaminyl transferase, gelatin liquefaction. The bacterium was positive for O-nitrophenyl-beta-D-galactopyranoside, N-acetylglucosaminyl transferase, gelatin liquefaction, and acid production from D-galactose, lactose, melibiose, raffinose, citrate, and trehalose. The bacterium was negative for indole production and acid production from maltose, α-methyl-D-glucoside, sorbitol, D-arabitol, inositol, inulin, and melezitose. All the strains exhibited pectolytic activity on potato slices. DNA from these strains yielded a single size amplicon with the primer set of PhF/PhR for P. wasabiae (De Boer et al. 2012) by PCR. However, DNA from these strains did not yield the expected amplicon with the primer set of BR1f/L1r for P. carotovorum subsp. brasiliense (Duarte et al. 2004) and Eca1f/Eca2r for P. atrosepticum (De Boer et al., 1995) by PCR. The sequence analysis of 16S rDNA (LC597917- LC597919) showed more than 98% identities to P. wasabiae strains (e.g. HAFL01 in Switzerland) by BLAST analysis. In addition, Multi-locus sequence analysis (Ma et al. 2007) was performed by MEGA6.06 using concatenated DNA sequences of seven housekeeping genes (aconitate hydratase(acnA, LC597925- LC597927), glyceraldehyde-3-phosphate dehydrogenase A(gapA, LC597972-LC597974), isocitrate dehydrogenase (icdA, LC597998- LC597998LC598000), malate dehydrogenase(mdh, LC598024- LC598026), mannitol-1-phosphate dehydrogenase (mtlD, LC598050- LC598052), glucose-6-phosphate isomerase (pgi, LC598076- LC598078) and gamma-glutamyl phospate reductase (proA, LC598099- LC598101)), and all clustered into a clade containing other confirmed strains of P. wasabiae. As a result, these three strains shared high identity with each other (>98%, E-Values showed 0). The clade containing these three strains was consistently placed in a larger clade with the other P. wasabiae and 100% bootstrap support for its separation from other Pectobacterium species available in GenBank when the consensus tree constructed using Maximum Likelihood method. Pathogenicity of these strains against onion (cv. 'Hayate') was confirmed by the field experiments with 5 weeks growth plants sprayed with bacterial suspension (1×107cfu/ml) resulting in soft rot on the plants about four weeks after inoculation, whereas water-inoculated plants remained symptomless. Strains re-isolated from the artificially diseased stems were confirmed as P. wasabiae using the methods as biochemical characterization and multiple genetic analyses. Based on the disease symptoms, the cultural, molecular, and pathological features of the strains, we conclude that the soft rot symptoms of onion in Hokkaido in 2019 were caused by P. wasabiae. To our knowledge, this is the first report of P. wasabiae as the soft rot disease agent of onion in Japan.

Effects of Ursolic Acid on Intestinal Health and Gut Bacteria Antibiotic Resistance in Mice.

Ursolic acid (UA), a natural pentacyclic triterpenoid, has been widely reported to exert anti-oxidant and anti-inflammatory properties. However, the effects of UA on the intestinal homeostasis and gut microbiota were rarely explored. The aim of the present study was to investigate the effects of UA on intestinal health and gut microflora antibiotic-resistance in antibiotic-exposed mice. Kunming mice (n = 80) were randomly allocated into three groups and fed with one of the following diets, respectively: Cont group (n = 20), the basal diet; UA group (n = 20), the basal diet supplemented with 150 mg/kg UA; Tet group (n = 40), the basal diet supplemented with 659 mg/kg chlortetracycline. After 14 days, 10 mice in each group were euthanatized and the remaining 30 mice in the Tet group were randomly allocated into three sub-groups (n = 10 per group) as follows: the Tet group which were kept feeding a Tet diet for 14 days; the Natural Restoration (NatR) group which received a basal diet for 14 days; and the UA therapy (UaT) group which fed a basal diet supplemented with 150 mg/kg UA for 14 days. Throughout the experiment, the weight and the food intake of each mouse were recorded once weekly. Serum LPS and diamine oxidase (DAO), jejunal morphology, jejunal tight junction proteins and nutrient transporters, colonic inflammatory cytokines, gut microbiota and its antibiotic resistance gene (ARG) were examined at euthanasia. The results showed that UA treatment significantly increased average daily food intake (ADFI) of mice. Notably, UA increased the jejunal villi height, decreased the jejunal crypt depth and promoted the expression of jejunum nutrient transporters. UaT group had higher villi height, lower crypt depth and higher nutrient transporter mRNA expression in jejunum than NatR group. Besides, UA decreased serum DAO content, upregulated mRNA expression of ZO-1, claudin-1 and occludin and downregulated TNF-α and IL-6. The mRNA abundances of ZO-1, claudin-1 and occludin and TNF-α and IL-6 in UaT group were, respectively upregulated and downregulated than NatR group. Furthermore, an analysis of 16S rDNA sequences demonstrated that UA increased the abundance of beneficial bacteria in the gut. And the results of ARG test showed that UA downregulated the expression of antibiotic-induced resistance genes. The UaT group inhibited the increase of harmful bacteria abundance and suppressed the mRNA abundances of ARG compared to the NatR group. In conclusion, considering the positive effects of UA on the growth performance and intestinal mucosal barrier, we anticipate that these findings could be a stepping stone for developing UA as a novel substitute of antibiotics.

Characterisation of a carotenoid producing extremely halophilic archaeon Halorubrum sodomense MS5.1 isolated from a solar saltern in Tamil Nadu, South India

A carotenoid producing extremely halophilic archaeon designated MS5.1 was isolated out of brine samples from a crystalliser pond of a marine solar saltern in Kanyakumari District, Tamil Nadu, South India. The red pigmented, long rod shaped haloarchaeon was found to be able to grow at temperature range of 20-40°C, salt concentration of 10-35% and pH range of 6 to 9 with optimum conditions for growth being 28°C; 30% salt and pH 7. The archaeal cells were found to be Gram negative and got lysed when placed in distilled water. Analysis of 16S rDNA sequence revealed that the isolate is phylogenetically related to species of the genus Halorubrum under the family Halobacteriaceae, with close relationship to Halorubrum sodomense. Further analyses of the phenotypic and biochemical characteristics of the isolate confirmed the identity of the organism as Halorubrum sodemense. The gene sequence of the strain was deposited in the NCBI GenBank with Accession No. MW332265. Polar lipid characterisation of the strain by thin layer chromatography (TLC) identified the major polar lipids as Phosphatidylglycerol (PG), Phosphatidylglycerol phosphate methyl ester (PGP-Me), Diglycosyl archaeol (DGA) and Sulfated diglycosyl archaeol (S-DGA). The strain was further screened for antibiotic sensitivity and found insensitive to antibiotics that target peptidoglycan layer and found sensitive only to Nitrofurantoin and Rifampicin, which works by inhibiting nucleic acid synthesis. As halophilic archaea are known natural sources of carotenoids, an attempt was made to extract these pigments from the cells and analysed by UV-VIS spectroscopy. Present study characterised the haloarchaeal strain H. sodomense MS5.1 isolated from a coastal solar saltern, optimised the growth conditions and the results clearly indicated that the strain is a potential source of carotenoids and halophilic enzymes.

Morphological descriptions of the nymph and adults of Ornithodoros clarki, the larva and nymph of Ornithodoros rondoniensis, with notes on their phylogenetic relationships.

Based on tick specimens collected recently in Mexico, Nicaragua, Panama and Brazil, we provide morphological descriptions of the nymph and adults of Ornithodoros clarki Jones & Clifford, 1972 from the first three countries, and the larva and nymph of Ornithodoros rondoniensis (Labruna, Terassini, Camargo, Brandão, Ribeiro & Estrada-Peña, 2008) from Brazil. Also, an analysis of mitochondrial 16S rDNA sequences was performed to analyze the phylogenetic relationships of these tick species. Adults and nymphs of O. clarki and O. rondoniensis are unique among the Argasidae family by presenting exceptionally large spiracular plates with small goblets, and an integument with smooth polygonal mammillae. However, these two species are morphologically distinct based on specific patterns of coxal folds, idiosomal mammillae and pilosity, and female genital flap. In contrast, the larvae of O. clarki and O. rondoniensis are morphologically identical, except for a general larger size of the former species; this slight difference is corroborated by Principal Component Analysis (PCA) by using 40 morphometric variables. Phylogenetic analyses including 16S rDNA partial sequences of different Ornithodoros taxa from Central and South America indicate that O. rondoniensis from Brazil diverges from O. clarki from Mexico, Nicaragua and Panama. However, phylogenetic distance separating both alleged species is similar or slightly lower than the distances depicted for conspecific populations of a few other Ornithodoros species. Nonetheless, our primary criterion to maintain O. rondoniensis as a valid species is because its adult and nymphal stages do present distinct morphological traits that easily distinguish these postlarval stages from O. clarki.

Characterization of Erwinia sp. causing black rot of papaya (Carica papaya) first recorded in Okinawa Main Island, Japan

AbstractSince 2002, papaya black rot has been spreading over several islands of Okinawa Prefecture, Japan. To devise a prevention strategy for the disease, microbiological research on the pathogen was conducted. Twelve strains were isolated from papaya infected with black rot showing symptoms such as water‐soaked lesions on stems and petioles, black spots on fruits, and rotted leaves turning yellow with necrotic spots. Through Koch's postulates, we confirmed that the isolated strains caused papaya black rot. Bacteriological assays showed that the strains have characteristics different from the type strains of Erwinia mallotivora, E. papayae, and E. psidii. Moreover, 16S rDNA sequence similarity searches showed that the isolated strains had less than 98.6% similarity with type strains. Additionally, phylogenetic analysis of 16S rDNA sequences suggested that the isolated strains were possibly a novel species belonging to the genus Erwinia, as the strains formed an independent cluster and had low sequence similarity with the type strains. Earlier studies indicated that papaya black rot is caused by E. cypripedii. Therefore, we propose to add the Erwinia sp. isolated in this study to the list of papaya black rot pathogens.

Molecular detection of phytoplasmas in witches’ broom diseased Ziziphus oenoplia from Andhra Pradesh, India

Ziziphus oenoplia is a wild jujube evergreen small tropical fruit tree that grows along the roadsides. During a survey in 2021, this jujube trees exhibited severe witches’ broom symptoms, therefore, an investigation was carried out, to verify the presence of phytoplasmas in the symptomatic plants. Total DNA was extracted from symptomatic leaves using a CTAB method and a PCR assay was performed using 16S rRNA gene specific primer pairs P1/P7 and R16F2n/R16R2. Amplicons of about 1.2 kb were consistently amplified from all the symptomatic plant samples in nested PCR assays, while no amplification was observed in non-symptomatic samples. Virtual RFLP analysis of 16S rDNA sequences allowed the classification of this strain into 16SrV group. The phylogenetic analysis of 16Sr DNA sequence of the detected phytoplasma strain indicates its close clustering with ‘Candidatus Phytoplasma balanitae’ and ‘Ca. P. ziziphi’ and further analyses on other housekeeping gene are necessary to define its affiliation.

Molecular identification of a ‘Candidatus Phytoplasma australasia’ strainin radish showing a little leaf disease in Andhra Pradesh, India

Symptoms of little leaf, proliferation of axillary buds and pale green colour leaves were observed in radish plants grown in the farmer fields at Anantapuram, Andhra Pradesh. DNA was extracted from symptomatic and asymptomatic leaves and subjected to nested PCR assay with P1/P7 and R16F2n/R16R2 primers. The about 1,250 bp amplicon was obtained only from symptomatic radish plants and the positive control, but not from any asymptomatic plants. Pair wise sequence comparison and phylogenetic analyses of 16S rRNA gene sequences of the radish phytoplasma strain suggested that it is related to ‘Candidatus Phytoplasma australasia’. Further virtual RFLP analysis of 16S rDNA sequences allowed the classification of the radish phytoplasma strain in the 16SrII-D subgroup that results to be in great expansion in this area of India.

Molecular identification of phytoplasmas associated with little leaf disease of brinjal in Andhra Pradesh and Telangana states of India

Brinjal little leaf (BLL) is a very important phytoplasma disease in India that causes severe economic losses every year. During 2020, symptoms of little leaf and witches’ broom were observed in brinjal plants grown in the farmer fields at Kadapa, Andhra Pradesh and Mahabubnagar district, Telangana states. Four symptomatic and two symptomless plants collected from the surveyed places of both states were subjected to DNA extraction and used as template in nested PCR assays with universal phytoplasma 16S rRNA primers. Amplification of about 1.25 kb product with primers R16F2n/R2 was obtained only from symptomatic BLL plants and the positive control, but not from asymptomatic plants. Pair wise sequence comparison and phylogenetic analyses of 16S rRNA gene sequences of these phytoplasma strains confirmed that the phytoplasmas from Mahabubnagar district in Telangana were ‘Candidatus Phytoplasma trifolii’-related strains, while those from Kadapa, Andhra Pradesh, were related to Ca. P australasia’. Further virtual RFLP analysis of 16S rDNA sequences allowed the classification of these strains into 16SrVI-D and 16SrII-D subgroups, respectively.

Molecular identification of 16SrI-X subgroup phytoplasma associatedwith eggplant in India

Eggplants exhibiting phyllody and virescence symptoms typical of phytoplasma infection were observed in fields in Alandi, Pune, Maharashtra, India. The phytoplasma presence was confirmed in all symptomatic samples using PCR and nested PCR with specific primer pairs P1/P7 and R16F2n/R2. Nucleotide sequence comparison revealed that the amplicon sequence shares more than 99% identity with ‘Candidatus Phytoplasma asteris’ sequences and in the phylogenetic analysis clustered with sequences of similar strains of phytoplasmas. Virtual RFLP analysis of 16S rDNA sequences enabled to enclose the strain into the 16SrI-X subgroup. This is the first report of natural occurrence of a 16SrI-X phytoplasma in eggplant.

ZFP90 drives the initiation of colitis-associated colorectal cancer via a microbiota-dependent strategy

ABSTRACT Chronic inflammation and gut microbiota dysbiosis are risk factors for colorectal cancer. In clinical practice, patients with inflammatory bowel disease (IBD) have a greatly increased risk of developing colitis-associated colorectal cancer (CAC). However, the underlying mechanism of the initiation of CAC remains unknown. Systematic analyses using an existing genome-wide association study (GWAS) and conditional deletion of Zfp90 (encoding zinc finger protein 90 homolog) in a CAC mouse model indicated that Zfp90 is a putative oncogene in CAC development. Strikingly, depletion of the gut microbiota eliminated the tumorigenic effect of Zfp90 in the CAC mouse model. Moreover, fecal microbiota transplantation demonstrated that Zfp90 promoted CAC dependent on the gut microbiota. Analysis of 16s rDNA sequences in fecal specimens from the CAC mouse model allowed us to speculate that a Prevotella copri-defined microbiota might mediate the oncogenic role of Zfp90 in the development of CAC. Mechanistic studies revealed Zfp90 accelerated CAC development through the TLR4-PI3K-AKT-NF-κB pathway. Our findings revealed the crucial role of the Zfp90-microbiota-NF-κB axis in creating a tumor-promoting environment and suggested therapeutic targets for CAC prevention and treatment.

Detection and identification of a phytoplasma associated with rapeseedfasciation in Iran

During surveys in 2017 and 2018, a disease was observed in rapeseed fields in east of Iran. Affected plants exhibited extensive fasciation, formation of bunches of flowers, lack of leaves formation and seed pod abortion. The symptomatic rapeseed plants were analyzed for phytoplasma presence by nested PCR using phytoplasma universal primers and all tested symptomatic rapeseeds plants were positive for phytoplasma presence, while asymptomatic rapeseed plants were negative. Nested PCR positive samples were directly sequenced and in silico RFLP and phylogenetic analyses of PCR-amplified 16S rDNA sequences indicated the presence of a phytoplasma enclosed in the peanut witches’ broom broom group (16SrII). This is the first report of phytoplasma presence in rapeseed plants showing fasciation.

Environment and host species identity shape gut microbiota diversity in sympatric herbivorous mammals.

The previous studies have reported that the mammalian gut microbiota is a physiological consequence; nonetheless, the factors influencing its composition and function remain unclear. In this study, to evaluate the contributions of the host and environment to the gut microbiota, we conducted a sequencing analysis of 16S rDNA and shotgun metagenomic DNA from plateau pikas and yaks, two sympatric herbivorous mammals, and further compared the sequences in summer and winter. The results revealed that both pikas and yaks harboured considerably more distinct communities between summer and winter. We detected the over-representation of Verrucomicrobia and Proteobacteria in pikas, and Archaea and Bacteroidetes in yaks. Firmicutes and Actinobacteria, associated with energy-efficient acquisition, significantly enriched in winter. The diversity of the microbial community was determined by the interactive effects between the host and season. Metagenomic analysis revealed that methane-metabolism-related pathway of yaks was significantly enriched in summer, while some pathogenic pathways were more abundant in pikas. Both pikas and yaks had a higher capacity for lipid degradation in winter. Pika and yak shared more OTUs when food shortage occurred in winter, and this caused a convergence in gut microbial composition and function. From winter to summer, the network module number increased from one to five in pikas, which was different in yaks. Our study demonstrates that the host is a dominant factor in shaping the microbial communities and that seasonality promotes divergence or convergence based on dietary quality across host species identity.

First report on the association of ‘Candidatus Phytoplasma asteris’ with lethal wilt disease of coconut (Cocos nucifera L.) in India

Lethal wilt disease of coconut, reported as a disease of unknown etiology as early as 2007, was for the first time found consistently in association with a phytoplasma in diseased coconut trees in Thanjavur, Thiruvarur and Pudukkottai districts in Tamil Nadu on the southeastern coast of India. Nested PCR with phytoplasma-specific universal primers and in silico restriction analysis of 16S rDNA sequences revealed the association of ‘Candidatus Phytoplasma asteris’ (16SrI-B) group phytoplasma, which was confirmed by sequence analysis of translation elongation factor (tuf) and ribosomal protein (rp) genes. Because this disease kills affected palms within 5 months, recommendations are given to arrest the spread of the pathogen to other coconut-growing regions in the country.

Exogenous dopamine and overexpression of the dopamine synthase gene MdTYDC alleviated apple replant disease.

Apple replant disease (ARD) is a soil-borne disease that leads to economic losses due to reduced plant growth and diminished fruit yields. Dopamine is involved in interactions between plants and pathogens. However, it remains unclear whether dopamine can directly stimulate defense responses to ARD. In this study, an exogenous dopamine treatment and dopamine synthetase MdTYDC (tyrosine decarboxylase) transgenic plants were used to verify the role of dopamine in treating ARD. First, 2-year-old apple trees (Malus domestica cv. Fuji), grafted onto rootstock M26, were grown in replant soils. The addition of dopamine (100μM) to the soil promoted seedling growth and changed the accumulation of mineral elements in plants in replant soils. Such supplementation improved the activity of invertase, urease, proteinase and phosphatase under replant conditions. Sequencing analysis of 16S rDNA and internal transcribed spacer (ITS) rDNA revealed that dopamine had a slight influence on bacterial diversity but had an obvious effect on the fungal diversity in replant soils. The application of dopamine to replant soil changed the composition of bacterial and fungal communities. Second, overexpression of MdTYDC in apple plants alleviated the effects of ARD. MdTYDC transgenic lines exhibited mitigated ARD through inhibited degradation of photosynthetic pigment, maintaining the stability of photosystems I and II and improving the antioxidant system. Furthermore, overexpression of MdTYDC improved arbuscular mycorrhizal fungi colonization by improving the accumulation of soluble sugars under replant conditions. Together, these results demonstrated that dopamine enhances the tolerance of apples to ARD.