Diversity and Localization of Bacterial Endosymbionts from Whitefly Species Collected in Brazil

Whiteflies (Hemiptera: Aleyrodidae) are sap-sucking insect pests, and some cause serious damage in agricultural crops by direct feeding and by transmitting plant viruses. Whiteflies maintain close associations with bacterial endosymbionts that can significantly influence their biology. All whitefly species harbor a primary endosymbiont, and a diverse array of secondary endosymbionts. In this study, we surveyed 34 whitefly populations collected from the states of Sao Paulo, Bahia, Minas Gerais and Parana in Brazil, for species identification and for infection with secondary endosymbionts. Sequencing the mitochondrial Cytochrome Oxidase I gene revealed the existence of five whitefly species: The sweetpotato whitefly Bemisia tabaci B biotype (recently termed Middle East-Asia Minor 1 or MEAM1), the greenhouse whitefly Trialeurodes vaporariorum, B. tabaci A biotype (recently termed New World 2 or NW2) collected only from Euphorbia, the Acacia whitefly Tetraleurodes acaciae and Bemisia tuberculata both were detected only on cassava. Sequencing rRNA genes showed that Hamiltonella and Rickettsia were highly prevalent in all MEAM1 populations, while Cardinium was close to fixation in only three populations. Surprisingly, some MEAM1 individuals and one NW2 population were infected with Fritschea. Arsenopnohus was the only endosymbiont detected in T. vaporariorum. In T. acaciae and B. tuberculata populations collected from cassava, Wolbachia was fixed in B. tuberculata and was highly prevalent in T. acaciae. Interestingly, while B. tuberculata was additionally infected with Arsenophonus, T. acaciae was infected with Cardinium and Fritschea. Fluorescence in situ hybridization analysis on representative individuals showed that Hamiltonella, Arsenopnohus and Fritschea were localized inside the bacteriome, Cardinium and Wolbachia exhibited dual localization patterns inside and outside the bacteriome, and Rickettsia showed strict localization outside the bacteriome. This study is the first survey of whitely populations collected in Brazil, and provides further insights into the complexity of infection with secondary endosymionts in whiteflies.     

Bacterial Infection and Immune Responses in Lutzomyia longipalpis Sand Fly Larvae Midgut

The midgut microbial community in insect vectors of disease is crucial for an effective immune response against infection with various human and animal pathogens. Depending on the aspects of their development, insects can acquire microbes present in soil, water, and plants. Sand flies are major vectors of leishmaniasis, and shown to harbor a wide variety of Gram-negative and Gram-positive bacteria. Sand fly larval stages acquire microorganisms from the soil, and the abundance and distribution of these microorganisms may vary depending on the sand fly species or the breeding site. Here, we assess the distribution of two bacteria commonly found within the gut of sand flies, Pantoea agglomerans and Bacillus subtilis. We demonstrate that these bacteria are able to differentially infect the larval digestive tract, and regulate the immune response in sand fly larvae. Moreover, bacterial distribution, and likely the ability to colonize the gut, is driven, at least in part, by a gradient of pH present in the gut.     

Bacterial and Fungal Diversity of Quaternary Cave Sediment Deposits

The bacterial and fungal assemblages of clastic sediments collected from two caves located in north-western Romania were investigated by assessing ITS and 16S rRNA gene diversity. Bacterial members belonging to Chloroflexi, Nitrospirae, Proteobacteria, Firmicutes, Acidobacteria, Gemmatimonadetes, and fungal members of Ascomycota were identified. Except for Bacillus sp., all bacteria were related to uncultured or unknown species and the majority (86%) of the bacterial sequences from one of the caves had no close GenBank relatives. The bacterial sequences obtained clustered with species found in extreme environments. Half of the bacterial operational taxonomic units were clustered with clones isolated from deep subsurface sediments of a radioactively contaminated site in the USA. The present study represents the first attempt to identify microorganisms in Quaternary cave sediments.

Supplemental materials are available for this article. Go to the publisher's online edition of Geomicrobiology Journal to view the supplemental file.     

The Diversity of Yellow-Related Proteins in Sand Flies (Diptera: Psychodidae)

Yellow-related proteins (YRPs) present in sand fly saliva act as affinity binders of bioamines, and help the fly to complete a bloodmeal by scavenging the physiological signals of damaged cells. They are also the main antigens in sand fly saliva and their recombinant form is used as a marker of host exposure to sand flies. Moreover, several salivary proteins and plasmids coding these proteins induce strong immune response in hosts bitten by sand flies and are being used to design protecting vaccines against Leishmania parasites. In this study, thirty two 3D models of different yellow-related proteins from thirteen sand fly species of two genera were constructed based on the known protein structure from Lutzomyia longipalpis. We also studied evolutionary relationships among species based on protein sequences as well as sequence and structural variability of their ligand-binding site. All of these 33 sand fly YRPs shared a similar structure, including a unique tunnel that connects the ligand-binding site with the solvent by two independent paths. However, intraspecific modifications found among these proteins affects the charges of the entrances to the tunnel, the length of the tunnel and its hydrophobicity. We suggest that these structural and sequential differences influence the ligand-binding abilities of these proteins and provide sand flies with a greater number of YRP paralogs with more nuanced answers to bioamines. All these characteristics allow us to better evaluate these proteins with respect to their potential use as part of anti-Leishmania vaccines or as an antigen to measure host exposure to sand flies.     

Long-Term Phosphorus Fertilization Impacts Soil Fungal and Bacterial Diversity but not AM Fungal Community in Alfalfa

Soil function may be affected by cropping practices impacting the soil microbial community. The effect of different phosphorus (P) fertilization rates (0, 20, or 40 kg P₂O₅ ha⁻¹) on soil microbial diversity was studied in 8-year-old alfalfa monocultures. The hypothesis that P fertilization modifies soil microbial community was tested using denaturing gradient gel electrophoresis and phospholipids fatty acid (PLFA) profiling to describe soil bacteria, fungi, and arbuscular mycorrhizal (AM) fungi diversity. Soil parameters related to fertility (soil phosphate flux, soluble P, moisture, phosphatase and dehydrogenase assays, and carbon and nitrogen content of the light fraction of soil organic matter) were also monitored and related to soil microbial ribotype profiles. Change in soil P fertility with the application of fertilizer had no effect on crop yield in 8 years, but on the year of this study was associated with shifts in the composition of fungal and bacterial communities without affecting their richness, as evidenced by the absence of effect on the average number of ribotypes detected. However, variation in soil P level created by a history of differential fertilization did not significantly influence AM fungi ribotype assemblages nor AM fungi biomass measured with the PLFA 16:1ω5. Fertilization increased P flux and soil soluble P level but reduced soil moisture and soil microbial activity, as revealed by dehydrogenase assay. Results suggest that soil P fertility management could influence soil processes involving soil microorganisms. Seasonal variations were also recorded in microbial activity, soil soluble P level as well as in the abundance of specific bacterial and fungal PLFA indicators of soil microbial biomass.     

Impact of Ectomycorrhizosphere on the Functional Diversity of Soil Bacterial and Fungal Communities from a Forest Stand in Relation to Nutrient Mobilization Processes

The ectomycorrhizal symbiosis alters the physicochemical and biological conditions in the surrounding soil, thus creating a particular environment called ectomycorrhizosphere, which selects microbial communities suspected to play a role in gross production and nutrient cycling. To assess the ectomycorrhizosphere effect on the structure of microbial communities potentially involved in the mobilization of nutrients from the soil minerals in a poor-nutrient environment, we compared the functional diversity of soil and ectomycorrhizosphere bacterial communities in a forest stand. Two hundred and sixty-four bacterial strains and 107 fungal strains were isolated from the bulk soil of an oak (Quercus petraea) stand and from oak–Scleroderma citrinum ectomycorrhizosphere and ectomycorrhizae, in two soil organo-mineral horizons (0 to 3 cm and 5 to 10 cm). They were characterized using two in vitro tests related to their capacities to mobilize iron and phosphorus. We demonstrated that the oak–S. citrinum ectomycorrhizosphere significantly structures the culturable bacterial communities in the two soil horizons by selecting very efficient strains for phosphorus and iron mobilization. This effect was also observed on the diversity of the phosphate-solubilizing fungal communities in the lower soil horizon. A previous study already demonstrated that Laccaria bicolor–Douglas fir ectomycorrhizosphere structures the functional diversity of Pseudomonas fluorescens population in a forest nursery soil. Comparing to it, our work highlights the consistency of the mycorrhizosphere effect on the functional diversity of bacterial and fungal communities in relation to the mineral weathering process, no matter the fungal symbiont, the age and species of the host tree, or the environment (nursery vs forest). We also demonstrated that the intensity of phosphorus and iron mobilization by the ectomycorrhizosphere bacteria isolated from the lower soil horizon was significantly higher compared to that which was isolated from the upper horizon. This reveals for the first time a stratification of the functional diversity of the culturable soil bacterial communities as related to phosphorus and iron mobilization.     

Robustness of the Bacterial Community in the Cabbage White Butterfly Larval Midgut

Microbial communities typically vary in composition and structure over space and time. Little is known about the inherent characteristics of communities that govern various drivers of these changes, such as random variation, changes in response to perturbation, or susceptibility to invasion. In this study, we use 16S ribosomal RNA gene sequences to describe variation among bacterial communities in the midguts of cabbage white butterfly (Pieris rapae) larvae and examine the influence of community structure on susceptibility to invasion. We compared communities in larvae experiencing the same conditions at different times (temporal variation) or fed different diets (perturbation). The most highly represented phylum was Proteobacteria, which was present in all midgut communities. The observed species richness ranged from six to 15, and the most abundant members affiliated with the genera Methylobacteria, Asaia, Acinetobacter, Enterobacter, and Pantoea. Individual larvae subjected to the same conditions at the same time harbored communities that were highly similar in structure and membership, whereas the communities observed within larval populations changed with diet and over time. In addition, structural changes due to perturbation coincided with enhanced susceptibility to invasion by Enterobacter sp. NAB3R and Pantoea stewartii CWB600, suggesting that resistance to invasion is in part governed by community structure. These findings along with the observed conservation of membership at the phylum level, variation in structure and membership at lower taxonomic levels, and its relative simplicity make the cabbage white butterfly larval community an attractive model for studying community dynamics and robustness.     

Diversity of Bacterial Isolates from Commercial and Homemade Composts

The diversity of heterotrophic bacterial isolates of three commercial and two homemade composts was studied. The commercial composts were produced from poultry litter (PC), sewage sludge (SC), municipal solid waste (MC), and homemade composts (thermal compost [DC] and vermicompost [VC]) from food wastes. The taxonomic and physiological diversity of the heterotrophic culturable bacteria was assessed using phenotypic and genotypic characterization and the analysis of the partial 16S rRNA gene sequence. Composts DC and SC presented the higher genotypic diversity, as could be inferred from the number of distinct genotypic patterns observed, 28 and 21, respectively. Gram-positive bacteria, mainly Firmicutes, were predominant in all the composts. Some organisms related with taxa rarely reported in composts, as Rhodanobacter spathiphylli, Moraxella osloensis, Lysobacter, Corynebacterium, Pigmentiphaga kullae, and new taxa were also isolated. The highest relative proportion of isolates able to degrade starch was found in compost SC (>70%), to degrade gelatine in compost DC (>70%), to degrade Tween 80 in compost PC (>90%), and to degrade poly-epsilon-caprolactones in compost DC (>80%). Compost MC presented the lowest relative proportions of isolates able to degrade starch (<25%), gelatine (<20%), and poly-epsilon-caprolactone (<40%). When compared with the others, the homemade composts presented higher relative proportions of Gram-positive isolates able to inhibit the target organisms Staphylococcus aureus, Enterococcus faecalis, Escherichia coli, or Pseudomonas aeruginosa. In compost MC, none of the Gram-positive isolates was able to inhibit those targets.     

抗生素对老年前期及老年患者所致肠菌群紊乱的探讨

本文对23例联合用广谱抗生素的老年前期及老年患者发生腹泻后,进行了肠菌群分析,部分做了难辨梭菌的培养。结果表明,全部被检患者应用广谱抗生素后都发生腹泻,应用抗生素种类繁多,其中以氨苄加庆大或先锋加庆大最多。取患者新鲜便作肠菌群分析,6例曾进行纤维结肠镜检查,其中2例肠粘膜有小片状伪膜附着。在使用大量、多种抗生素后,最短在半天内,大多在2周内即发生腹泻,菌群中肠杆菌、拟杆菌、双歧杆菌、乳杆菌,肠球菌、葡萄球菌、酵母菌、小梭菌及真杆菌有显著的差异,均明显低于正常,6/15例难辨梭菌为阳性,检出率40％。治疗先停用抗生素加强支持疗法或用健康人粪便滤液灌肠,15例停止腹泻,2例使用万古霉素才停止。     

Microbial Diversity in Sediments Collected from the Deepest Cold-Seep Area, the Japan Trench

The Japan Trench land slope at a depth of 6,400 m is the deepest cold-seep environment with Calyptogena communities. Sediment samples from inside and beside the Calyptogena communities were collected, and the microbial diversity in the sediment samples was studied by molecular phylogenetic techniques. From DNA extracted directly from the sediment samples, 16S rDNAs were amplified by the polymerase chain reaction method. The sequences of the amplified 16S rDNAs selected by restriction fragment length polymorphism analysis were determined and compared with sequences in DNA databases. The results showed that 33 different bacterial 16S rDNA sequences from the two samples analyzed fell into similar phylogenetic categories, the α-, γ-, δ-, and ɛ-subdivisions of Proteobacteria, Cytophaga, and gram-positive bacteria; some of the 16S rDNA sequences were common to both samples. δ- and ɛ-Proteobacteria-related sequences were abundant in both sediments. These sequences are mostly related to sulfate-reducing or sulfur-reducing bacteria and epibionts, respectively. Eight different archaeal 16S rDNA sequences were cloned from the sediments. The majority of the archaeal 16S rDNA sequences clustered in Crenarchaeota and showed high similarities to marine group I archaeal rDNA. A Methanococcoides burtonii–related sequence obtained from the sediment clustered in the Euryarchaeota indicating that M. burtonii–related strains in the area of Calyptogena communities may contribute to production of methane in this environment. From these results, we propose a possible model of sulfur circulation within the microbial community and that of Calyptogena clams in the cold-seep environment. Received June 15, 1998; accepted November 10, 1998.     

Bacterial contamination of amniotic membrane in a tissue bank from Iran

Human Amniotic Membrane (AM) transplantation can promote tissue healing and reduce inflammation, tissue scarring and neovascularization. Homa Peyvand Tamin (HPT) tissue bank has focused on manufacturing human cell and tissue based products including AM. The purpose of this study is to evaluate and identify bacterial contamination of AMs that is produced by HPT for several ophthalmic applications. From July 2006 to April 2011, 122 placentas from cesarean sections were retrieved by HPT after obtaining informed consent from the donors. Besides testing donor’s blood sample for viral markers, microbiological evaluation was performed pre and post processing. During tissue processing, decontamination was performed by an antibiotic cocktail including; Gentamicin, Ceftriaxone and Cloxacillin. Of 271 cesarean section AM donors who were screened as potential donors, 122 were accepted for processing and assessed for microbiological contamination. Donors’ age were between 21 and 41 years (Mean = 27.61 ± 0.24). More than 92 % of mothers were in their first or second gravidity with full term pregnancies. The most prevalent organisms were Staphylococci species (72.53 %). After processing, contamination rates markedly decreased by 84.62 % (p value = 0.013). According to our results, most of bacterial contaminations were related to donation process and the contamination pattern suggests procurement team as a source. Therefore we recommend that regular training programs should be implemented by tissue banks for procurement staff. These programs should focus on improved donor screening and proper aseptic technique for tissue retrieval. We also suggest that tissue banks should periodically check the rate and types of tissue contaminations. These data help them to find system faults and to update processing methods.     

Mineral and Bacterial Diversities of Desert Sand Grains from South-East Morocco

Mineralogy and microbiology of sand from Merzouga (Morocco) were simultaneously characterized, with the purpose of contributing to a better understanding of the geomicrobiology of deserts. In spite of very low measured bacterial biomass, bacterial diversity on each of the five defined mineralogical classes, was found high. An original grain by grain cultivation method enabled to obtain bacterial isolates with an unusually high recovery rate. The results of this study show that the genus Arthrobacter is well adapted to this environment with a preference for grains other than the dominant mineral quartz, and that the genera Chelatococcus and Saccharotrix are strongly attached to the grains.     

山羊瘤胃微生物宏基因组BAC文库的构建与分析

从山羊瘤胃液中提取混合微生物DNA,经BamHI部分酶切得到50kb～800kb的DNA片段后,将其连接到pCCIBAC载体上,转化E．coliEPI300,建立山羊瘤胃微生物BAC文库。经RFLP鉴定分析,该文库12672个克隆,平均插入片段为6lkb。该文库的构建为后续新型基因的筛选提供了材料,为进一步研究山羊瘤胃微生物奠定了基础。     

Isolation and Characterization of Bacterial Species from Ain Mud Volcano,Iran

Along the coastal belt of Makran, on the southeastern margin of Iran, there are several active and inactive mud volcanoes with different morphologies. Ain—an actively bubbling eye-shaped mud pool—is one of these unique geological phenomena. Present study indicates the first overview of the bacterial diversity of this mud volcano obtained by culture techniques. For this purpose, two samples were collected at two different depths of the mud pool and were cultivated on two different nutrient culture media. A total of 13 isolates were randomly chosen for further phenotypic and genotypic characterization. Growth of the isolates occurred at 25–42°C, pH 8–10 and 0–10% NaCl, indicating that most of them were haloalkaliphiles. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the bacterial isolates belonged to three major taxa: Gammaproteobacteria (genera: Marinobacter, Nitrincola, Stentrophomonas), Actinobactera (genera: Kocuria, Brevibacterium, Dietzia) and Firmicutes (genus Planomicrobium). Gammaproteobacteria were the most abundant, following Actinobacteria and Firmicutes. Most of the strains isolated in the present study had the ability to produce extracellular hydrolytic enzymes such as lipases, amylases, proteases and DNases, making them important biotechnologically.     

Opportunistic Fungal Infections in the Epidemic Area of COVID-19: A Clinical and Diagnostic Perspective from Iran

Mycopathologia - The coronavirus disease 2019 (COVID-19) pandemic emerged in Wuhan, China, in late 2109, and has rapidly spread around the world. Until May 25, 2020, there were 133,521 confirmed...     

Sequencing-Based Analysis of the Bacterial and Fungal Composition of Kefir Grains and Milks from Multiple Sources

Kefir is a fermented milk-based beverage to which a number of health-promoting properties have been attributed. The microbes responsible for the fermentation of milk to produce kefir consist of a complex association of bacteria and yeasts, bound within a polysaccharide matrix, known as the kefir grain. The consistency of this microbial population, and that present in the resultant beverage, has been the subject of a number of previous, almost exclusively culture-based, studies which have indicated differences depending on geographical location and culture conditions. However, culture-based identification studies are limited by virtue of only detecting species with the ability to grow on the specific medium used and thus culture-independent, molecular-based techniques offer the potential for a more comprehensive analysis of such communities. Here we describe a detailed investigation of the microbial population, both bacterial and fungal, of kefir, using high-throughput sequencing to analyse 25 kefir milks and associated grains sourced from 8 geographically distinct regions. This is the first occasion that this technology has been employed to investigate the fungal component of these populations or to reveal the microbial composition of such an extensive number of kefir grains or milks. As a result several genera and species not previously identified in kefir were revealed. Our analysis shows that the bacterial populations in kefir are dominated by 2 phyla, the Firmicutes and the Proteobacteria. It was also established that the fungal populations of kefir were dominated by the genera Kazachstania, Kluyveromyces and Naumovozyma, but that a variable sub-dominant population also exists.