Questions related to Marine Microbiology
I am trying to grow marine cyanobacteria from my samples using BG11 and L1 as nutrient and agar as gelling agent. I found agarolytic bacteria thrives in my plates, liquifying my agar. So, I tried using gellan gum as an alternative to agar. In my trials, I keep getting wobbly unstable gellan gels even though I used magnesium sulphate as cations. After overnight upside-down, I got dome-shaped gellan gel surface. When I tried using cell spreader, the gelling broke. Has anyone encounter this problem?
I'm looking for an untouched area or highly emerged problem that humans (probably farmers, pharmaceutical industry-related) facing recently or might be faced in the near future which can be sorted out by marine microbiological or biotechnological resources.
or please suggest to me any resource related to this matter. your valuable comments are highly appriciated.
I want to isolate fungi from sea water. Which medium do you recommend to do so?
I was thinking about PDA modified with sea water but I am not sure if I should use either 100% sea water or a mixture distilled or deionized water/sea water? I do not know either if it is better to use natural filtered sea water or to prepare artificial sea water.
Thank you in advance
While extracting MPs from sediment samples of Peruvian sandy beaches, I found a transparent film with dark brown dots (see pictures attached). Could those be colonies of marine microorganisms? If so, what are the possible species or families? Consider this sample coming from Lima, Peru (SE Pacific).
I'm interested in studying a Master Degree program on Marine Microbiology or related fields.
I wanted to know if you can recommend me a place or pogram, based on your experience.
Thank you all for your time.
I am attempting to culture aerobic marine methanotrophs and was planning on plating them on seawater agar. I keep finding references to 'aged' seawater in media recipes and was wondering what the necessity of aging the seawater in the dark is? If the aging is necessary, should I filter sterilize or autoclave the seawater prior to aging or just before media prep?
Thanks for your input!
The composition of the BH medium is as follows:
MgSO4 - 0.2g/L
CaCl2 - 0.02g/L
KH2PO4 - 1g/L
K2HPO4 - 1g/L
(NH4)2SO4 - 1g/L
and FeCl3 - 0.05g/L
I am conducting a comparative study trying to isolate bacteria on different concentrations of Marine broth (MB) media + agar vs MB + gellan gum/Gelzan/Gelrite.
For the agar I have 10, 50, and 100% MB, which solidifies perfectly fine.
I have tried 10, 50, and 100% MB + gellan gum but only 10% MB solidifies. I have tried to optimise the concentrations of gellan gum from the recommended 6.5 g/L to 10 g/L. I have also tried to prepare the media with and without Mg2SO4.
Any advice would be appreciated!
In order to gather knowledge regarding state of the art tag sequencing methods used in the field of microbial oceanography, I am looking for on going projects of marine microbial observatories/recently published studies regarding time series of marine microbial communities.
Thanks in advance!
i am more interest to know what kind of hypothesis, question, or objective that each index usually answer. surprisingly that both indices has quite the same definition in a term of scribing the reason of using them to define diversity. i calculate both indices and i would like to discuss the results. simply my objective is to see if there are special and temporal differences in phytoplankton diversity among 8 sampling stations.
also i am interested to know what are the main objectives or questions of your researches that you answered using such indices. thanks
I have been isolating Vibrio spp. from sea food using TCBS agar, but have a lot of confusion in morphological identity and biochemical tests. Most of the suspected isolates were identified as Aeromonas and Shewanella after 16S rRNA sequencing.
I would be thankful if anyone can let me know about any other convenient method to identify Vibrio spp. and I need to know whether 16S rRNA sequencing is a reliable method to identify Vibrio.
I am in search of protocols for culturing marine bacteria from sea sediments under anoxic or microaerophilic conditions.
My samples are from deep-sea sediments 2-5 km in depth from Antarctic waters. The sediments are within the upper 10 cm of the seabed.
Any protocols, media suggestions, or advice would be extremely appreciated.
Hi, I am approaching to the identification of nano and microplankton using an inverted microscope with epifluorescence.
I would like to differentiate the organisms in 7 classes:
Autotrophic dinoflagellates, autotrophic flagellates, cryptophytes, diatoms, prymnesiophytes, heterotrophic dinoflagellates and
This type of classification has been used by Landry et al. 2014 (http://dx.doi.org/10.1016/j.dsr2.2014.02.006).
Using the epifluorescence I can distinguish heterotrophs from autotrophs, but I do not now how to identify the organisms for each class.
Do you know how to do it?
Thank you, any help will be more than welcome,
I am analysing microbial assemblages attached to marine microplastics using 16S metagenomics, and I have a query about maximising the quantity of DNA I can extract from the substrate?
I have been using the DNeasy Kit, Qiagen (Formerly PowerSoil, MoBio) to remove the attached assemblage from the plastic (recommended by Harrison et al, 2014), but I am struggling to extract any DNA - according to PCR amplification and 0.8% agarose gel electrophoresis. I am aware that the associated biomass, ergo DNA yield, is typically very low.
One paper (Debeljak et al, 2017) suggests that I should incorporate an additional lysis step using ReadyLyse solution (Epicentre) to maximise cell lysis. However, I was wondering if there was a cheaper alternative? For example, I was thinking of creating a Lysozyme:TE Buffer solution to the desired molarity, although I'm aware that the enzymes are derived from different sources and have different efficiencies.
If anyone has any feedback/recommendations, that would be very useful!
Thanks in advance.
Can fastidious or even unculturable marine bacteria be grown in situ? If so, what are the current methodologies for doing so?
I am looking for standard nutrient solid media recipes or protocols used to culture marine bacteria. What are the standard types of media used to culture marine bacteria (psychrophiles)?
Also, in scenarios where these media are not able to culture these bacteria what methods are then employed?
I am investigating marine bacteria for potential plastic degradation, and wanted to know if there any standardised protocols or selective media that would be worth considering during the screening process.
I have considered directly incorporating the polymer into the media (PE powder & marine agar - Lobelle and Cunliffe, 2011), Bushnell-Haas media (Harshvardhan & Jha, 2013) and the MATH assay. However, my understanding is the latter two methods typically investigate liquid hydrocarbon, whereas I am using a solid substrate.
If anyone has any suggestions, or is facing a similar issue, or has any feedback on the above mentioned methods, please comment
I work with soils and sediments from Antarctica, and as DNA is very stable under those low temperatures, I would like to use RNA extractions instead of DNA extractions from the environmental samples to do amplicon sequencing with Illumina. Is that possible? have anyone did amplicon sequencing for 16S, 18S, ITS, based on cDNA from RNA extractions?
Hi, we are working on stool-metagenomics based on V3V4 and ITS2 amplicon sequencing. We would like to analyse data in a work station by using QIIME. Can someone help me with the best configuration (type of processor, RAM, hard disk storage) of work station that is required for analysing this NGS data (preferably for the data that is generated on an Illumina-MISeq platform; assuming for each sample nearly 2 GB of data is generated; and we need to analyse a maximum of about 120 such samples at a time).
I am presently carrying out research on microalgae that produces toxin in lagos Nigeria, i would like to know where i can purchase saxitoxin standard or where i can send my samples to (water and shellfishes) for quantification. Thanks in advance .
This is a recurring unknown in my samples. It is about 20 um in length. The sample was preserved with Lugol's solution, and the image was taken from an inverted microscope. The sample was taken from a central Texas reservoir in June.
I want to culture the tardigrades collected from intertidal area? In the initial experiment, I tried with diatom feed(Achnanthes) but failed.
I have isolated Botryococcus in a river, when I analysed it through LM it has colonies and mucilage threads connecting. But when I tried culturing it became unicellular but there are some visible threads, but it has a thick cup shape mucilage on the outer cell and takes 14 days to culture while optimising its culture conditions. But later on ill do some ecological studies and phylogenetic studies on that river. But my theory is that the botryococus I have isolated is not in river itself but the source of it at top or upstream. I hope you can help me.
I am landlocked an unable to culture marine specimens myself. I am having trouble locating an institution which has this culture available for study.
I want to quantify bacteria present both in the tissue and skeleton of coral and am thinking of inoculating 96 well plates and doing serial dilutions to obtain the most probable number (MPN). I can use a plate reader to measure optical density so human error is removed when assessing turbidity of the wells. This poses the question:
1) How can I remove a standardised area, weight or volume of coral tissue and skeleton with minimal damage to the coral?
I need this to standardise the number of bacteria to i.e. number of bacteria per ml mucus or per cm2 tissue. I've tried using a scalpel to scrape or break off pieces but it is difficult to standardise this and I've tried using 25G needles to extract tissue from the polyps, though this is only successful on long-polyped morphologies. I've heard that Waterpik-ing is very crude and difficult to standardise too.
Any insight on how to quantify coral-associated bacteria would be helpful. I had also thought about qPCR but this too requires removing a standardised area of coral tissue and skeleton to begin with.
I was planning to run a qPCR assay on amoA genes (bacterial and archaeal) from my marine sediment DNA extracts. After going through literature I observed that many people were using the Arch-amoAF and Arch-amoAR to amplify archaeal amoA (Francis et al 2005) and amoAF1 and amoA2R for the bacteria (Rotthauwe et al 1997). However these are the same primers used for preparing standards for their assays. I have a pure culture N.maritimus to be used as a standard for the archaeal assay. I was wondering if I could use the above mentioned primer to amplify the amoA gene from the cell culture, purify the product and directly use it as a standard without cloning it ? My main concern was if it was fine to run the qPCR assay with primers that were used to prepare the standard itself.
My lysmata seticaudata have become sterile, they are still carrying eggs as normal, and these are developing but not hatching, and are then shed with the moult. Their eyes have a growth covering them, can anyone identify this, is it likely to be related to the sterility issue? Lysmata boggessi have also become sterile, but I've moved them on before I'd noticed this eye issue. Attached are two photos, i4 is a healthy eye, i5 and i8 have the growth.
Does anybody know that can the Ceratium furca and Ceratium fusus produce toxic？Are they Harmful algae？THX！
I am currently studying for my masters and my project involves studying chytrids and their association with marine diatoms. I have a lot of papers many of which relate to previous methods and findings in freshwater environments. I was wondering if anyone had observed possible or confirmed chytrids in conjunction with any other research into diatoms and if so did you notice any patterns or species that seemed particularly affected. I'm not looking for definitive data just any thoughts or observations anyone may have made.
I am looking to determine the rate at which a seawater intake pipe will foul from biological material at about 40 feet water depth off southern California
We are currently trying to create mutualistic associations between yeast and algae and our system relies on identifying a carbon source that is not utilised by Chlorella species. However, thus far it seems to be able to grow on most carbon sources that we tested.
This question came up after reading the paper by Engelhardt et al., 2015.
Water samples were taken from an estuary (Salinity: 15 ppt) and brown colony formed in the agar plates after 4 weeks of plating. Photographs taken at 40 X magnification. Size of the cells: 8-15 micron
I am looking for a specific microbe and though someone here could help? For my research, I need a non-pathogenic bacterial host that is slightly halophilic. Ideally this host should be able to survive in fresh or seawater and preferably one who's genome has been sequenced in the past.
I am currently working with E. coli NEB 10-b and it is proving to be very sensitive to changes in ionic strength. I am also working with P. putida KT2440. Though I have managed to grow this host in LB media with an ionic strength of 0.5 M, they cannot survive the change to glucose minimal media at the same ionic strength (let alone surviving in seawater).
Does anyone have a suggestion or potential host in mind?
I have several aerobic marine bacteria strains that I would like to test for sulfide resistance. My plan would be to add Na2S to the medium (marine broth), but as far as I know the S will precipitate with the oxygen, which is not the point.
After looking online and in few books I could not find a solution.
Anybody has an idea?
I isolated many phages from marine water and took a microscopic image for this phages but now I have many phages. I think this may be new phage but I'm not sure whether these are new phages or not. Also I every time I take microscopic images for phages isolated from one plaque, I see more than one phage in my sample.
I read many articles about purification of phages from marine water. I did all that was advised but I don't get good results. Thank you for advise about all of these issues.
I want to obtain the cyanobacterium "Synechococcus sp. MA19" to carry out experiments.
Can you provide me with a source, e.g. a culture collection? I contacted authors that describe this strain (papers from 2000-2001) and large culture collections, but so far I was not able to find/obtain a sample MA 19.
Originally, the MA 19 was isolated from a hot spring in Japan (Miyakejima).
I am looking for a multiparametric probe instead of a set of probes for specific nutrients, e.g. nitrate, phophates and silicates.
ex: I have isolated Streptomyces albus from marine sediments. can I name it some thing like M1 or S1. Any standard systematic codes are there to name the strain?
I have many phage isolated from marine environment and I want to measure their Capsid and tail size . please if you have program for this issue please share it to me.
I am currently using the SILVA reference database and I get OTUs assigned to MALV III. I saw in Georges et al. 2014 they were observed in Southern Ocean, but find little about their relation to the rest of the MALV. Anyone has papers or hits to help me?
I have initiated my work in Cyanobacteria from metagenomic cyanobacterial samples, and i wish to do some taxonomy work at molecular level. I am using 16S rRNA gene based Cynano bacterial primers CY106F and CY 781 R (Nubel et al., 1997). During PCR i am getting nonspecific bands along with the primers. I have tried all things by changing annealing temperature, MgCl2, BSA . But cant able to solve the problem. My query is is there any other Cyanospecific primers available? I had seen some people were using 16S rRNA and ITS based primers for Cyano classification. I am quite optimistic with 16S rRNA gene based classification. And also how fare morphology based classification will help compared to molecular level? Comments and suggestions are welcome!!!
I want to know if bacteria requires insoluble hydrocarbon as a sole carbon source for biosurfactant production or they can produce it during their normal growth without any insoluble hydrocarbon.
I will be going on a 2-3 day trip and was wondering if I could just isolate bacteria (from sediments) in the field instead of bringing the samples back to the lab. If I have to bring the samples back then I would have to shorten my trip. Are there any Dos and Don'ts for this? or anything to take heed off?
People usually make glycerol stock and store at -80 freezer. Do you remove salts before making the glycerol stock?
I'd like to get a Trichodesmium thiebautii starter culture and so far, I've only found T. erythraeum. Does anybody have an idea where/which organization I could ask? I'd be extremely grateful for any help!
I am trying to get the standards for Mycosporine like amino acids to charaterize for the same in cyanobacteria but it is not commercially available. Please suggest me in this regard.
Adults and eggs of killifish kept in a small modular recirculation system, showed fungal disease (cotton like appearance) on the anal fin and around the egg corion.
I am going to investigate the growth of Celeribacter spp. ( isolated from marine environment) at different salt concentration by using Luria Bertani broth. I was wondering if anybody could tell me, what other ingredients except trypton, yeast extract, NaCl should be added to LB?
I am looking to run survival experiment of bacteria in a lake mesocosm and would like to count viable cell numbers over time in sediments. Ideally, I would like to be able to calculate CFU/ml and not CFU/g so I can compare to growth in lake water as well. I have tried Beat Beating with 2mm beads but my recovery is very low. Any suggestions or ways people get around the mass to volume comparisons!?
Why I did not get good digestion for phage DNA? I need to determine the diversity of phage isolates, and the approximate genome size from the fragments generated from RFLP. My concentration of phage DNA is 338.97 ng/µl & 1.93 at 260/280 nm. I used 1µl EcoR1 (NEB), 1µl (50x) buffer, 5µl DNA and 3µl Mill Q water for 10 µl reaction.
I’m using specific primers to quantify Scrippsiella donghaienis (a phytoplankton species) in DNA extracts from sediments by real time PCR. When total DNA concentration is high, the melting curve shows a specific peak of Scrippsiella donghaienis at 82,5 °C. However, in sediments of low DNA concentration, I get a double peak (a very small peak of Scrippsiella and another higher peak at 88 °C). Do you have an explanation to this ? I attached 2 images of specific amplification and non specific amplification from the same test.
I'm looking for papers/tutorials/information on how to classify transcripts based on species. For example, if I was going to be sequencing a small anemone doing a whole-body RNA extraction I want to be able to differentiate between the anemone and the food it has eaten (bacteria, plankton, etc.).
I know that the Lehnert et al. 2014 (Extensive Differences in Gene Expression between Symbiotic and Aposymbiotic Cnidarians) designed a custom Support Vector Machine algorithm "TopSort"; however, it is not open-source.
If anyone could provide other examples (preferably with open-source software that would require no, or very little, tweaking) It would be most appreciated. I think that gut microbiome or metagenomic research would cover this but I haven't found anything yet.
Matthew J. Oldach
In the culture and screening of genetically modified microalgae, which can endure against the antibiotics, cells maintained duplication but green color of microalgae slowly disappears (not perfectly). What does it mean?