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Welcome to your first training module for becoming a PMN volunteer. 

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This is a self guided "tour" of the basic information required to participate 

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and you can refer back to this recording at any time.  

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We will be covering three main topics: 

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1) what exactly harmful algal blooms are, 

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2) some information about our phytoplankton monitoring network,

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and 3) how you will be collecting samples in the field 

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and analyzing your water samples 

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for the presence of particular types of phytoplankton,

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which we refer to as our "target organisms." 

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The P in PMN stands for phytoplankton, 

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which comes from the Greek words for plant and wanderer. 

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Many of them can photosynthesize just like plants on land 

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and they are at the mercy of the movements of the currents and winds.

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Phytoplankton is a general term 

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used to describe single-celled, photosynthetic microscopic organisms that live in aquatic environments. 

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Also called micro algae, 

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they live in mainly in the water column 

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where they can get enough sunlight to produce their own food, 

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thus forming the basis of the aquatic food chain. 

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The oxygen produced 

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is released into the water and the atmosphere, 

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which provides about half the oxygen that we breathe. 

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Phytoplankton are also important in carbon cycling 

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and climate control of the planet. 

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So life truly would not exist without phytoplankton. 

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It is important to keep these crucial benefits in mind 

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as we will be concentrating only on those

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that can potentially cause harmful algal blooms.

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Harmful algal blooms, or HABs, 

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can occur when phytoplankton grow at very rapid rates. 

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This can lead to obvious discoloration of the water 

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on small and large scales. 

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We prefer to use the term HAB 

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rather than “red tide” or “brown tide” 

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because there are a variety of colors a HAB can be, 

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and can even be harmful without discoloration of the water, 

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depending on the species of phytoplankton present. 

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Some can even bioluminesce. 

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Phytoplankton growth is generally seasonal, 

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and depends on the availability of 

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carbon dioxide, sunlight, and nutrients. 

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Phytoplankton, like land plants, require nutrients 

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such as nitrate, phosphate, silicate, and calcium 

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at various levels depending on the species. 

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Other factors influence phytoplankton growth rates, 

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including water temperature and salinity, 

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water depth, wind, 

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and what kinds of predators are grazing on them.

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When excess nutrients are available 

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under ideal growth conditions, 

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blooms are more likely to appear.

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Research is being done to better predict when blooms may form

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to help mitigate their potential harmful effects, 

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but not all algal blooms are harmful! 

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For example, a phytoplankton called Coccolithophorids 

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does not produce a toxin and are generally harmless, 

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even though they can form blooms so large 

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that you can see them from space. 

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And there were so many in the ancient ocean 

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that as they died and sank to the bottom

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they formed chalky sediments layers 

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thick enough for you to walk on top of today, 

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like at the Cliffs of Dover. 

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So what do we mean when we say "harmful"?

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There are several ways 

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in which a bloom can be considered harmful.

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One is if they cause physical damage. 

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For example, fish gills can become clogged 

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or damaged by the phytoplankton cells. 

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Fish respond to this damage by producing mucus, 

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and this can ultimately lead to death by suffocation. 

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When a bloom dies off, 

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the cells sink through the water column 

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and can be degraded by bacteria. 

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This process removes oxygen from the water, 

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making survival impossible for other organisms 

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due to the formation of dead zones 

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from hypoxic or anoxic conditions.

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What most people often associate with a harmful algal bloom

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is the production of toxins. 

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There are many different groups of toxins 

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that vary in their levels of toxicity, 

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and health effects can be seen in marine animals 

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like dolphins and sea turtles, as well as in humans. 

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As seen on the map, this is a global issue, 

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with some regions susceptible to multiple types of toxin exposure.

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Not all phytoplankton are able to produce toxins; 

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in fact, very few are. 

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They must have the genes for toxin production, 

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and even if they HAVE the genes, 

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those genes may not always be turned on. 

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But if toxins are being made, they can travel through the food web 

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and concentrate in the tissues of things like shellfish and fish. 

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Exposure comes from either feeding on the toxic cells directly or 

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feeding on something ELSE that has been exposed to the toxins.

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Depending on the amount of toxin ingested, 

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effects can range from relatively mild to lethal. 

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Bloom-forming toxic microalgae include three main groups of phytoplankton: 

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the dinoflagellates, the diatoms, and cyanobacteria.

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These three types of algae produce the toxins 

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which cause most human and animal health problems.

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The known toxin-producing species 

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typically do not affect human health directly, 

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but illness occurs through the consumption 

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of toxin-contaminated seafood or drinking water. 

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That said, sometimes toxic species don’t produce any toxin, 

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and the amount of toxin, again, can vary among species, as well. 

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These are the five human health syndromes 

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that are known to be directly attributed to algal toxin exposure. 

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The primary causative agent is shown in each, 

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although, in some cases, 

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it is not necessarily the only organism capable

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of producing the different groups of toxins, shown in red.

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Symptoms will vary depending on

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the type of toxin that has been ingested and how much, 

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but in some instances, such as with saxitoxins,

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it can be lethal in humans. 

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Unfortunately, it is not as simple as just looking at a bloom 

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and being able to determine if it is toxic 

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just by the organisms present or by discoloration of the water. 

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Toxins can be present even if the water is not discolored.

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This could occur if the cells have lysed,

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opened upon death,

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and the toxins were released into the water,

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but there is no color change.

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Another way is if the bloom is deep enough in the water, 

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such that it is not visible from the surface. 

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As previously mentioned, 

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not all species have the genes to produce toxins, 

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and if they do, they are not always turned on. 

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So, the presence of a species known to produce toxins 

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does not always mean that toxins are present.  

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And for each type of toxin, there are multiple forms or congeners, 

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each with varying degrees of toxicity.

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All of these confounding factors 

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make it more difficult to immediately tell if a bloom is toxic or not, 

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but knowing the potential for risk 

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based on identification of what is in the sample 

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is still extremely beneficial information to have 

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from a monitoring standpoint. 

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Because of the potential harmful effects of HABs, 

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monitoring for the presence of the organisms 

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that may lead to them is important for several reasons. 

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We know they are increasing in how often they happen, 

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they are beginning earlier in the year than before 

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and lasting longer, 

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and they are also expanding 

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in their geographic range of occurrence.

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By knowing which organisms are present 

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and in what relative abundance, 

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we can use that information with other tools, like satellite imagery,

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to better understand and predict where HABs might occur. 

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Routine monitoring can also allow us to know early on

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when a bloom might be starting to form   

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and thus act as an early warning system 

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to reduce potential exposure to toxins.

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So, why help PMN?

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Most agencies lack the funding and resources 

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to monitor the entire coastline of a state, 

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or look at every lake and reservoir that can be impacted by HABs. 

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Having local citizens engaged in this effort 

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by being the “boots on the ground” 

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is crucial to increasing the capacity of monitoring 

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and better understanding where and when HABs occur, 

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thus helping NOAA and other scientists in their HAB research.

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PMN is a national volunteer organization that is

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trained to monitor for potentially harmful algae or cyanobacteria. 

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You will be trained on how to collect samples  

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from coastal or freshwater environments, 

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how to identify the organisms specific to your region, 

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and how to enter that data into NOAA’s database.

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Based on what our volunteers find, 

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we can test for toxins (in a limited capacity), 

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alert the state or local agency

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that is responsible for making management decisions about HABs 

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and look at long-term trends 

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of how HABs may be changing in how often and where they occur. 

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We have volunteers from a wide variety of

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backgrounds and groups, both science and non-science

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and our volunteers have generated 

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over 300,000 data points across the country since our inception. 

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PMN began here in Charleston, South Carolina, 

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back in 2001 with coastal monitoring, 

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and in 2015 we expanded into freshwater HAB monitoring 

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thanks to an interagency agreement with EPA’s Office of Water. 

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As a PMN volunteer, 

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you will be provided with all of the equipment seen here. 

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The only equipment we can not provide is a microscope, 

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but everything else needed to collect water samples, 

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measure salinity and temperature, 

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make slides and ship samples to us will be sent to you. 

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There is also an app called Phyto 

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that you can download if you have an iPhone or iPad. 

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This is another useful resource

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to help with the identification of what’s in your sample. 

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There are photos of both coastal and freshwater organisms, 

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pronunciations of the scientific names, 

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and a key based on cell shape 

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to help narrow down your identification choices. 

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Unfortunately, the Android version is still being updated, 

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but the original, older Android version 

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that lacks the freshwater information may still be available. 

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The second part of the training will concentrate on teaching you 

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how to identify the target organisms for your region. 

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You will learn how to identify to the genus level of classification, 

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as the features needed to get to the species level are difficult, 

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or impossible to see with a regular light microscope. 

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If we need to determine 

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if you have a toxin-producing species present, 

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you will send us the sample and we will use our 

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powerful scanning electron microscope to make that determination.

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Feedback will be provided, 

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so you will know if you have correctly identified your organisms.

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This feedback is an important part of our program, 

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both to create a strong working relationship 

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between volunteers and NOAA staff, 

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as well as a way to ensure the quality of the data generated. 

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And our volunteers produce results. 

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They have been able to identify many blooms over the years, 

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and some of those were found to be toxin-producing blooms 

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that the state regulatory agencies were not aware of. 

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As previously mentioned, many blooms are harmless, but knowing

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which ones have the potential to be harmful is very important!  

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Even our newer freshwater program 

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has found toxin-producing blooms. 

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The data generated by our volunteers is not just for PMN usage, 

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but other entities use our data as well,

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for building models for things like drought and tide gauges 

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to recreational fisheries trying to avoid potential HAB areas. 

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This portion will cover the protocols 

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you will be using to collect samples 

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and how to go through that sample on your microscope. 

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You should have already viewed the training video. 

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that covers many of these steps.

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If not, please email pmn@noaa.gov to receive a copy. 

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The first thing to do 

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is select the site that you will be sampling on a regular basis.

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Where you sample is completely up to you, 

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but you need to be able to get there consistently, 

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at least every other week for a year. 

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It could be a dock, along a beach, or at a park, 

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wherever you can safely, and legally, tow your plankton net. 

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You need to be able to get there, again, on a consistent basis.

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For picking your site,

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this could be someplace where you have seen blooms,

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or discolored water before;

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someplace that's important to you or your community,

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like a neighborhood pond, or beach;

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or you might just be curious to see what's in your water.

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You don't necessarily need to sample

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on the exact same day every week, or at the same time.

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You can schedule according to what works best for you.

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But, again, you do need to be sampling at least every other week.

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Once you decide what site you are going to use,

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your homework is to send us your site information. 

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00:13:53,314 --> 00:13:55,233
This would include a name for your site

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00:13:55,587 --> 00:13:58,970
and GPS coordinates, ideally in decimal format. 

259
00:13:59,728 --> 00:14:03,566
This allows us to get your site information into our data entry portal 

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00:14:03,970 --> 00:14:05,637
and make you a dot on the map. 

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00:14:06,394 --> 00:14:08,415
It is possible to have more than one site, 

262
00:14:08,768 --> 00:14:11,142
so we would need the information for each site.

263
00:14:14,323 --> 00:14:17,707
Here is an overview of what PMN sampling and analysis involves. 

264
00:14:18,010 --> 00:14:20,535
We will go through the field sampling steps briefly, 

265
00:14:20,535 --> 00:14:22,556
as they were covered in the training video.

266
00:14:25,838 --> 00:14:29,172
You will receive a copy of a region-specific data sheet. 

267
00:14:29,879 --> 00:14:33,767
This shows the target organisms you are looking for in particular. 

268
00:14:34,424 --> 00:14:37,505
You are not responsible for identifying everything you see in your 

269
00:14:37,505 --> 00:14:41,646
water sample, but you are responsible for being able to identify

270
00:14:41,646 --> 00:14:42,808
the target organsims, 

271
00:14:43,111 --> 00:14:46,141
as these have the potential to cause harmful algal blooms.

272
00:14:47,303 --> 00:14:49,373
You should bring a copy of this data sheet with you 

273
00:14:49,373 --> 00:14:52,252
each time you sample, in order to record your data. 

274
00:14:53,211 --> 00:14:53,767
The date, 

275
00:14:54,323 --> 00:14:55,434
time of sampling, 

276
00:14:55,838 --> 00:14:58,262
water and air temperatures, along with salinity, 

277
00:14:58,262 --> 00:15:01,141
are all required information for each sample. 

278
00:15:02,252 --> 00:15:05,787
Items below the dotted line on the data sheet are all optional, 

279
00:15:05,787 --> 00:15:09,272
like weather conditions and other measurements you can take 

280
00:15:09,373 --> 00:15:11,191
if you have the equipment to make them. 

281
00:15:11,999 --> 00:15:15,029
We will cover more details about the data sheet shortly. 

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00:15:19,625 --> 00:15:22,352
So, first, you are going to do your plankton tow.

283
00:15:22,706 --> 00:15:25,938
It is important that there is no air in your net or the bottle. 

284
00:15:26,544 --> 00:15:29,170
You'll dunk your net until the bottle is full of water.

285
00:15:29,574 --> 00:15:32,251
Then, let it sink vertically as deep as you can

286
00:15:32,251 --> 00:15:33,918
without hitting the bottom. 

287
00:15:34,372 --> 00:15:37,806
This depth can vary with tides and could be several feet, 

288
00:15:38,109 --> 00:15:40,635
or just a few inches deeper than the mouth of the net.

289
00:15:41,948 --> 00:15:43,311
After your vertical drop, 

290
00:15:43,311 --> 00:15:46,998
you'll bring the net back up to about a foot below the surface,

291
00:15:47,352 --> 00:15:51,442
and you'll begin your three-minute timer for the horizontal tow.

292
00:15:53,008 --> 00:15:55,836
Try to avoid getting anything in the net that is not water, 

293
00:15:55,836 --> 00:15:57,503
like sea grass or trash, 

294
00:15:57,907 --> 00:16:00,836
and avoid towing nets through visible oil or gas slicks.

295
00:16:01,897 --> 00:16:04,877
Do not let the net bump into pilings that may have mussels 

296
00:16:04,877 --> 00:16:07,503
or barnacle growth, as this may tear the net. 

297
00:16:08,513 --> 00:16:11,493
Please refer to the training video for more details on the net tow 

298
00:16:11,745 --> 00:16:14,270
and how to concentrate your sample at the end. 

299
00:16:16,846 --> 00:16:19,826
You will also need to record air and water temperatures. 

300
00:16:20,583 --> 00:16:22,603
You can tie a line on to the thermometer 

301
00:16:22,957 --> 00:16:25,179
and tie it off to a cleat or railing 

302
00:16:25,432 --> 00:16:27,805
and throw it into the water while you do your net tow. 

303
00:16:28,411 --> 00:16:29,775
You do want to give the thermometer 

304
00:16:29,775 --> 00:16:31,846
time to acclimate to the water temperature, 

305
00:16:32,351 --> 00:16:35,179
and take your reading quickly after removal from the water.

306
00:16:36,239 --> 00:16:38,310
Do not let the thermometer lay on the ground or dock 

307
00:16:38,310 --> 00:16:41,037
when taking air temperature, as this will not be accurate.

308
00:16:41,694 --> 00:16:43,057
You can also tie it to a railing 

309
00:16:43,057 --> 00:16:45,078
to hang while you do your salinity reading, 

310
00:16:45,431 --> 00:16:47,552
or hold it at the top of the thermometer

311
00:16:47,704 --> 00:16:49,774
to get an accurate air temperature.

312
00:16:52,148 --> 00:16:55,885
You will also be using a refractometer to take a salinity reading.

313
00:16:56,542 --> 00:16:59,825
Be sure to have the platform flooded with water before closing

314
00:16:59,825 --> 00:17:03,259
the cover, and point it towards the brightest part of the sky 

315
00:17:03,259 --> 00:17:06,289
to get good contrast between the blue and white backgrounds. 

316
00:17:07,198 --> 00:17:09,774
You will take the reading where those two backgrounds meet,

317
00:17:10,229 --> 00:17:12,602
using the values on the right-hand side.

318
00:17:17,148 --> 00:17:19,774
The last thing to do is collect the whole water samples 

319
00:17:19,774 --> 00:17:23,208
in the 1 L and 30 mL bottles. 

320
00:17:24,117 --> 00:17:25,784
All you have to do is fill them up.

321
00:17:26,440 --> 00:17:27,501
While at your site,

322
00:17:27,551 --> 00:17:30,228
be sure to take a look around and observe the conditions,

323
00:17:30,632 --> 00:17:33,864
including, optional weather details as seen on the data sheet,

324
00:17:34,369 --> 00:17:37,551
whether the water is discolored or smells bad, etc.

325
00:17:38,864 --> 00:17:40,379
The entire sampling process 

326
00:17:40,379 --> 00:17:43,763
should take about 10–15 minutes total.

327
00:17:47,147 --> 00:17:48,763
You do want to take care of your equipment, 

328
00:17:48,915 --> 00:17:50,632
so please be sure to give everything 

329
00:17:50,632 --> 00:17:53,106
a good freshwater rinse when you return home. 

330
00:17:53,864 --> 00:17:55,632
Do not use soap on the plankton net 

331
00:17:55,632 --> 00:17:57,551
and be sure to hang it up to dry, 

332
00:17:57,551 --> 00:18:00,379
as opposed to letting it lie in a sink or a tub.

333
00:18:04,722 --> 00:18:07,955
Ideally, you want to look at your sample as soon as possible,

334
00:18:08,106 --> 00:18:10,934
and definitely within 6–8 hours of collection. 

335
00:18:11,944 --> 00:18:14,823
You will take your sample from the 125 mL bottle 

336
00:18:14,823 --> 00:18:16,591
that was attached to the net. 

337
00:18:17,651 --> 00:18:19,217
Gently re-suspend the bottle.

338
00:18:19,974 --> 00:18:21,944
Then, collect some water with your pipette, 

339
00:18:22,550 --> 00:18:25,732
and add TWO drops to your gridded slide.

340
00:18:26,843 --> 00:18:29,015
Place a cover slip on top, as shown, 

341
00:18:29,570 --> 00:18:33,610
placing one edge of the cover slip on top of the slide, and 

342
00:18:33,610 --> 00:18:37,247
lowering the opposite edge as low as you can before letting it go. 

343
00:18:38,055 --> 00:18:39,923
This will reduce the amount of air bubbles 

344
00:18:39,923 --> 00:18:41,439
trapped under the cover slip.

345
00:18:42,095 --> 00:18:43,661
Then, you’re ready to do your analysis.

346
00:18:47,045 --> 00:18:50,428
Each gridded slide is an 8x8 gridded square, 

347
00:18:50,630 --> 00:18:54,721
and you will be looking through all 64 grids for your analysis. 

348
00:18:55,782 --> 00:18:58,963
The objective is to look for any target organisms present 

349
00:18:59,418 --> 00:19:01,640
and determine the relative abundance of each.

350
00:19:02,751 --> 00:19:04,367
You can start in any corner, 

351
00:19:04,923 --> 00:19:07,095
using your 10x objective. 

352
00:19:08,004 --> 00:19:11,993
Move around in that grid, and look for things that might be targets. 

353
00:19:12,903 --> 00:19:16,943
If you see one, focus it in the exact middle of your field of view, 

354
00:19:17,599 --> 00:19:21,741
move up to your 40x objective, and make your identification.

355
00:19:23,054 --> 00:19:26,791
If it is a target organism, this is when you would take a photo. 

356
00:19:27,751 --> 00:19:31,690
Once you’ve made your ID then go back down to the 10X objective 

357
00:19:31,993 --> 00:19:33,710
and continue viewing through that grid 

358
00:19:33,710 --> 00:19:35,225
until it has been completely viewed.

359
00:19:36,185 --> 00:19:38,508
Then, move to the next grid, and begin again. 

360
00:19:39,215 --> 00:19:42,245
You will go through all 64 grids using this manner 

361
00:19:42,801 --> 00:19:46,387
and using a lawn-mower method, going row by row, 

362
00:19:46,639 --> 00:19:49,013
to ensure every square is viewed.

363
00:19:50,023 --> 00:19:52,851
You will not need to count all cells,

364
00:19:53,306 --> 00:19:54,972
but you do want to keep track 

365
00:19:54,972 --> 00:19:57,649
of how often you observe each target organism.

366
00:19:58,457 --> 00:20:00,427
This will be covered in more detail shortly.

367
00:20:04,113 --> 00:20:07,497
Taking photos of target organisms is crucial for our program.

368
00:20:08,103 --> 00:20:11,083
Any time you identify something as a target organism,

369
00:20:11,437 --> 00:20:14,063
we will need one or two representative photos,

370
00:20:14,315 --> 00:20:16,840
so that we can confirm your identification.

371
00:20:17,800 --> 00:20:20,174
This way we will ensure that the data that get entered 

372
00:20:20,174 --> 00:20:23,002
into our database are correct and are validated. 

373
00:20:24,063 --> 00:20:25,881
You will be given feedback on your IDs, 

374
00:20:25,881 --> 00:20:27,244
so you will know how you’re doing.

375
00:20:28,002 --> 00:20:32,093
We do not need a photo of EVERY single target cell you may see, 

376
00:20:32,446 --> 00:20:35,022
but, again, at least one or two representatives 

377
00:20:35,022 --> 00:20:38,052
of each target group that are present in your sample.

378
00:20:39,466 --> 00:20:43,456
Photos should be taken at 400X total magnification, 

379
00:20:44,062 --> 00:20:46,941
which is when you are using your 40X objective. 

380
00:20:47,900 --> 00:20:51,638
If you identify something that is a non-target organism, 

381
00:20:52,395 --> 00:20:54,567
meaning a more common type of phytoplankton

382
00:20:54,567 --> 00:20:56,284
that is not necessarily harmful,

383
00:20:56,991 --> 00:20:58,607
take a photograph of that, as well.

384
00:20:59,617 --> 00:21:01,385
If you are not sure what something is,

385
00:21:01,940 --> 00:21:04,365
take a photograph, and we will try to ID it for you.

386
00:21:05,274 --> 00:21:09,668
The goal is for YOU to make the ID, and for us to confirm it.

387
00:21:17,243 --> 00:21:18,657
The video I'm about to show

388
00:21:19,415 --> 00:21:22,092
lets you know how to use your cell phone camera

389
00:21:22,092 --> 00:21:24,263
to take photographs through a microscope.

390
00:21:25,425 --> 00:21:28,506
The most important part of this process is stability, 

391
00:21:28,506 --> 00:21:30,223
and it does take some practice.

392
00:21:50,323 --> 00:21:50,980
So,

393
00:21:53,404 --> 00:21:55,323
this is one method to take a photograph

394
00:21:55,323 --> 00:21:57,545
if you don't have a dedicated camera

395
00:21:57,545 --> 00:21:59,464
already connected to your microscope.

396
00:22:00,070 --> 00:22:02,444
Stability is the most important part of this process.

397
00:22:02,848 --> 00:22:05,424
So, using the three fingers on your left hand,

398
00:22:05,828 --> 00:22:07,343
brace your hand against the eyepiece.

399
00:22:08,404 --> 00:22:11,131
Hold your phone between your thumb and first finger. 

400
00:22:11,989 --> 00:22:14,161
Hold the other side with your opposite hand 

401
00:22:14,161 --> 00:22:17,090
so the thumb is free to click your camera button. 

402
00:22:17,999 --> 00:22:19,312
Starting a few inches away,

403
00:22:19,312 --> 00:22:22,242
find the center of the circle of white light with the camera 

404
00:22:22,595 --> 00:22:23,959
and move the phone closer 

405
00:22:23,959 --> 00:22:26,635
until the image in the microscope comes into focus. 

406
00:22:27,595 --> 00:22:29,565
Use your thumb to take the photograph. 

407
00:22:30,474 --> 00:22:31,989
To get rid of the black edges, 

408
00:22:31,989 --> 00:22:34,817
you can zoom in slightly first on your camera, 

409
00:22:35,474 --> 00:22:37,948
then, again, stabilize with your left hand,

410
00:22:38,605 --> 00:22:42,191
focus the white light until the object comes into focus 

411
00:22:42,544 --> 00:22:44,110
and there should be no black edge 

412
00:22:44,110 --> 00:22:45,423
when you take your photograph.

413
00:22:46,079 --> 00:22:48,302
This does take practice.

414
00:22:48,857 --> 00:22:49,766
So, keep trying! 

415
00:22:49,766 --> 00:22:52,847
Don't get frustrated if it doesn't work for you right away.

416
00:22:57,594 --> 00:22:59,716
You do not want to email your photos to us 

417
00:22:59,716 --> 00:23:01,534
directly from your cell phone

418
00:23:01,534 --> 00:23:04,160
if that is the method you use to take pictures.

419
00:23:04,816 --> 00:23:07,796
Please upload all photos to your computer first.

420
00:23:09,160 --> 00:23:11,079
When you're finished analyzing your slide,

421
00:23:11,079 --> 00:23:14,261
you're going to label each photo with your identification.

422
00:23:15,018 --> 00:23:16,735
Once all the photos are labeled, 

423
00:23:17,089 --> 00:23:20,574
you're going to complile all the photographs into one file.

424
00:23:21,129 --> 00:23:25,574
This can be a Word document, a PDF, or a PowerPoint presentation.

425
00:23:26,533 --> 00:23:30,725
Your PMN ID, your site name, and the sample date

426
00:23:30,725 --> 00:23:32,038
should also be included.

427
00:23:33,402 --> 00:23:35,624
This could be put into the file name,

428
00:23:36,129 --> 00:23:37,492
or at the beginning of the file, 

429
00:23:37,492 --> 00:23:39,614
as you see in the example on the right.

430
00:23:40,624 --> 00:23:44,058
You will email this file to PMN@noaa.gov

431
00:23:44,513 --> 00:23:47,088
and this is what you would receive feedback on.

432
00:23:48,553 --> 00:23:50,876
Please be patient with us to receive your feedback,

433
00:23:51,280 --> 00:23:52,947
but it is, usually, within one week.

434
00:23:53,553 --> 00:23:55,017
If you are just starting out, 

435
00:23:55,017 --> 00:23:58,704
please submit your photos before you enter your data

436
00:23:59,411 --> 00:24:02,744
to reduce corrections that may need to be made to the database.

437
00:24:03,653 --> 00:24:07,441
So, by submitting your photos first, and getting the feedback,

438
00:24:07,795 --> 00:24:10,168
you will be entering the corrected data.

439
00:24:16,178 --> 00:24:19,158
Again, you will receive a data sheet specific to your region,

440
00:24:19,158 --> 00:24:21,936
with your target organisms listed at the top.

441
00:24:24,815 --> 00:24:27,340
When you are done looking through the entire slide, 

442
00:24:27,845 --> 00:24:31,734
you need to have an answer of NO, YES, or ELEVATED 

443
00:24:32,138 --> 00:24:35,420
for each of the listed targets for your region.

444
00:24:37,592 --> 00:24:42,744
NO means that you did not see any of that particular target organism.

445
00:24:45,269 --> 00:24:47,844
YES means you did see at least one cell

446
00:24:47,895 --> 00:24:51,380
and maybe up to a few cells over the entirety of the slide.

447
00:24:54,410 --> 00:24:58,652
ELEVATED means that you see multiple cells of that target in a given grid, 

448
00:24:59,157 --> 00:25:01,329
and in a majority of the grids on the slide,

449
00:25:01,581 --> 00:25:03,450
meaning this is your dominant organism.

450
00:25:04,309 --> 00:25:07,036
Again, there’s no need to count the cells,

451
00:25:07,541 --> 00:25:10,218
but we are asking for a relative abundance.

452
00:25:10,925 --> 00:25:13,702
Are there NONE? Are there SOME? Are there LOTS?

453
00:25:14,712 --> 00:25:16,329
Here are some examples to show you.

454
00:25:20,369 --> 00:25:26,126
NO means exactly that: No cells for that target were seen anywhere on the slide.

455
00:25:28,702 --> 00:25:34,308
YES means that you are seeing some of that target scattered across your slide.

456
00:25:35,116 --> 00:25:39,106
For example, if you see just one Dinophysis, which is this cell here,

457
00:25:40,015 --> 00:25:42,490
and that is the only one you see, it is a YES.

458
00:25:43,449 --> 00:25:46,378
Now, if you see a few Dinophysis cells here and there,

459
00:25:46,833 --> 00:25:50,722
maybe some here, maybe some over there, one or two over there,

460
00:25:51,075 --> 00:25:56,681
but, overall, they are found in less than 60% of the 64 grids on your slide,

461
00:25:57,237 --> 00:25:58,550
then that is still a YES.

462
00:25:59,913 --> 00:26:04,105
Where to draw the line between a YES and an ELEVATED is subjective,

463
00:26:04,560 --> 00:26:07,035
but it does get easier with practice.

464
00:26:08,903 --> 00:26:12,640
Now, the lower end of ELEVATED might start to look something like this.

465
00:26:13,449 --> 00:26:15,570
In each of these grids,

466
00:26:15,873 --> 00:26:19,913
so we’re assuming each of these photos represents one grid on your slide,

467
00:26:20,519 --> 00:26:24,054
there are multiple cells of the same target organsim.

468
00:26:25,266 --> 00:26:31,630
If you start to see this in 60% or more of the grids overall,

469
00:26:31,832 --> 00:26:38,599
meaning you are seeing multiple cells of that target in the majority of your grid,

470
00:26:39,054 --> 00:26:42,236
then that would be considered a lot, or ELEVATED.

471
00:26:45,720 --> 00:26:47,589
Sometimes this is easier to determine.

472
00:26:48,094 --> 00:26:51,528
Here, you see a common phytoplankton called Rhizosolenia,

473
00:26:51,932 --> 00:26:53,549
which look like glass cigars.

474
00:26:54,811 --> 00:26:57,892
Rhizosolenia is dominating this particular grid, 

475
00:26:58,043 --> 00:27:00,316
and, presumably, the rest of the slide as well. 

476
00:27:01,276 --> 00:27:04,053
There are still other species of phytoplankton visible, 

477
00:27:04,356 --> 00:27:07,235
but it is clearly dominated by the Rhizosolenia.

478
00:27:07,841 --> 00:27:10,972
This would definitely fall into the elevated category.

479
00:27:12,639 --> 00:27:17,083
And if there is one species that seems to cover the entire slide,

480
00:27:17,538 --> 00:27:20,366
then that is most certainly elevated and a bloom.

481
00:27:21,073 --> 00:27:26,679
One usually sees discoloration of the water sample with concentrations of cells like those shown here.

482
00:27:29,406 --> 00:27:36,325
Hopefully, this gives you a visual idea of how to bin relative abundances during your sample analysis.

483
00:27:37,133 --> 00:27:42,386
Remember, you must have an answer of no, yes, or elevated for each target on your data sheet.

484
00:27:42,992 --> 00:27:46,527
These categories are not describing the sample as a whole.

485
00:27:51,123 --> 00:27:55,062
The last thing to do, after you have analyzed your slide, is to enter your data.

486
00:27:55,719 --> 00:27:58,295
This is another very important part of our program. 

487
00:27:58,850 --> 00:28:01,527
Once you provide a site name and GPS position,

488
00:28:01,830 --> 00:28:03,901
you’ll be given a PMN ID,

489
00:28:04,052 --> 00:28:06,173
which will correspond to the region that you are in.

490
00:28:07,284 --> 00:28:11,981
To enter your data, you’ll go to our homepage, and click the “submit data” button.

491
00:28:12,890 --> 00:28:15,062
You will then choose your desiganted region,

492
00:28:15,365 --> 00:28:17,688
and it will bring you to a Google form to fill out.

493
00:28:18,698 --> 00:28:20,516
You will type in your email,

494
00:28:21,324 --> 00:28:26,779
find your region, your PMN ID, and site name,

495
00:28:27,284 --> 00:28:30,617
and then fill in all the required information from your data sheet.

496
00:28:31,577 --> 00:28:35,162
For optional information, like the weather, or non-target organisms,

497
00:28:35,718 --> 00:28:38,799
those fields are pre-filled as “not applicable.”

498
00:28:39,455 --> 00:28:40,920
You may fill them in, if you would like.

499
00:28:42,283 --> 00:28:48,142
Photos are not uploaded here, but are emailed separately to PMN@noaa.gov

500
00:28:51,374 --> 00:28:53,344
So, what do you do with your bottles once you are done?

501
00:28:53,950 --> 00:28:56,475
That depends on what you have found in your sample.

502
00:28:57,233 --> 00:28:58,546
You’ll notice on your data sheet,

503
00:28:58,596 --> 00:29:02,031
that there are symbols in your “no,” “yes,” and “elevated” columns.

504
00:29:02,839 --> 00:29:08,293
These symbols will tell you which bottles you should preserve and save for shipment to us here at NOAA.

505
00:29:09,253 --> 00:29:12,990
The description of what each symbol means is found on your data sheet.

506
00:29:15,212 --> 00:29:19,909
We will first go over how to preserve samples, and then how often to ship them.

507
00:29:23,444 --> 00:29:25,515
If a symbol tells you to preserve a sample,

508
00:29:25,515 --> 00:29:29,757
you will add something called Lugol’s solution, which will also be provided to you.

509
00:29:30,414 --> 00:29:33,696
This kills the cells, but keeps them intact for further viewing. 

510
00:29:34,606 --> 00:29:39,605
This solution is harmless, but it will stain your skin or clothing, so please be careful when using it.

511
00:29:41,272 --> 00:29:45,211
If you are asked to preserve your 30-mL bottle,

512
00:29:45,514 --> 00:29:49,555
you will fill the provided pipette with Lugol’s up to the first mark,

513
00:29:55,060 --> 00:29:56,827
That is about 0.5 mL. and add that to the bottle.

514
00:29:57,938 --> 00:30:00,262
You will add that to your 30-mL bottle.

515
00:30:04,201 --> 00:30:08,039
If you are asked to preserve your 125-mL net tow bottle, 

516
00:30:08,595 --> 00:30:11,271
you will fill it to the 2nd mark with Lugol’s, 

517
00:30:11,574 --> 00:30:14,150
and add that to your 125-mL bottle.

518
00:30:15,261 --> 00:30:18,342
A preserved bottle should have the color of strong, iced tea.

519
00:30:19,150 --> 00:30:21,776
If your sample is very dense with phytoplankton, 

520
00:30:21,776 --> 00:30:25,261
you may need to add some additional Lugol’s to achieve that color.

521
00:30:26,423 --> 00:30:30,362
You will never add Lugol’s to the 1-Liter bottle!

522
00:30:32,887 --> 00:30:37,988
Once you’re done, you’ll add black, electrical tape to seal the lid shut,

523
00:30:38,594 --> 00:30:40,867
add a filled-out sample label,

524
00:30:41,725 --> 00:30:43,745
and put your bottles in a Ziploc baggie.

525
00:30:44,604 --> 00:30:48,493
You will want to store your preserved bottles and your Lugol’s solution

526
00:30:48,493 --> 00:30:51,169
in a cool, dark place—like a cabinet, or a drawer—

527
00:30:51,472 --> 00:30:55,210
until it’s time to ship the bottles to PMN, according to the calendar.

528
00:30:55,967 --> 00:30:57,886
We will go through some examples, now.

529
00:31:00,866 --> 00:31:03,038
So, if after looking through your entire slide

530
00:31:03,038 --> 00:31:07,684
you do not see any target organisms, you will have all NOs.

531
00:31:08,543 --> 00:31:10,108
The symbols are all circles,

532
00:31:10,916 --> 00:31:14,704
so you do not need to save ANY of your bottles for shipment.

533
00:31:15,361 --> 00:31:18,846
You will empty and rinse all three, and save them to reuse again.

534
00:31:19,805 --> 00:31:22,482
Remember, though, even if you do not see any targets,

535
00:31:22,936 --> 00:31:25,462
you still need to enter the data for that sample!

536
00:31:26,219 --> 00:31:29,047
It’s very important that you enter data for each and every sample,

537
00:31:29,047 --> 00:31:31,017
even if no targets are present.

538
00:31:34,502 --> 00:31:38,138
For this example, notice that there are circles in the YES column, as well.

539
00:31:38,795 --> 00:31:43,289
But, if at the end you still have all circles, the same applies:

540
00:31:43,643 --> 00:31:47,380
empty and reuse all three bottles. There is no need to save them.

541
00:31:50,613 --> 00:31:54,148
If you have a triangle in either the YES or ELEVATED column, 

542
00:31:54,552 --> 00:31:59,501
you will preserve both the 30-mL and the 125-mL net tow bottle. 

543
00:32:00,612 --> 00:32:03,289
Once they’re preserved, put them in your drawer, or cabinet, 

544
00:32:03,289 --> 00:32:06,471
and wait to ship them until the calendar says to.

545
00:32:07,632 --> 00:32:10,612
If you have any diamonds in the YES or ELEVATED column,

546
00:32:10,612 --> 00:32:13,036
you will only save the 30-mL bottle.

547
00:32:13,996 --> 00:32:17,733
Any bottle you are not asked to save, you will rinse and reuse.

548
00:32:18,491 --> 00:32:24,602
This means that 99.99% of the time you will be entering the 1-Liter bottle.

549
00:32:27,733 --> 00:32:29,955
You will be receiving a shipping calendar.

550
00:32:30,511 --> 00:32:36,470
This will tell you when to send all of your saved sample bottles over the course of two months.

551
00:32:37,531 --> 00:32:40,207
For example, if you start sampling in July,

552
00:32:40,561 --> 00:32:43,844
then you would send all of your preserved bottles in September,

553
00:32:44,046 --> 00:32:46,470
according to your region as noted at the bottom.

554
00:32:48,793 --> 00:32:51,672
You, again, will place the bottles in a Ziploc bag.

555
00:32:52,278 --> 00:32:57,076
You’ll place all of those in a white, plastic envelope that is provided,

556
00:32:57,682 --> 00:33:00,308
and you will use the appropriate shipping label.

557
00:33:00,813 --> 00:33:05,358
Most of the time, you will be using this brown, ground shipping label.

558
00:33:08,187 --> 00:33:12,580
You just need to fill out the areas marked in red with your information,

559
00:33:12,934 --> 00:33:17,580
and drop the package off at any UPS store, or call for UPS pick up.

560
00:33:18,439 --> 00:33:22,025
You should never be charged to ship your samples with these labels!

561
00:33:26,216 --> 00:33:30,459
Now, there may be a time when you have a square for your symbol.

562
00:33:31,317 --> 00:33:34,348
These are when you see elevated levels of a target

563
00:33:34,398 --> 00:33:39,095
that can potentially cause one of the five human health syndromes mentioned earlier.

564
00:33:39,953 --> 00:33:43,489
These are the samples that we want to have shipped IMMEDIATELY to us.

565
00:33:44,549 --> 00:33:46,468
You’ll preserve the two smaller bottles.

566
00:33:47,024 --> 00:33:49,650
You’ll, again, NOT add Lugol’s to the 1-Liter,

567
00:33:49,953 --> 00:33:52,226
but you will send the 1-Liter bottle, as well.

568
00:33:53,084 --> 00:33:56,115
And all three bottles need to be sent overnight.

569
00:33:58,337 --> 00:34:02,276
Only in this instance will you use the red, overnight UPS label. 

570
00:34:02,882 --> 00:34:04,953
You will fill it out the same as the ground label,

571
00:34:04,953 --> 00:34:08,539
but please remember to either take the “shipper’s” carbon copy,

572
00:34:09,094 --> 00:34:13,438
or take a photograph of the label, so you have a copy of the tracking number.

573
00:34:14,650 --> 00:34:17,730
You will email us to let us know a sample is coming overnight,

574
00:34:17,882 --> 00:34:20,306
so we can prepare to work it up upon arrival.

575
00:34:20,508 --> 00:34:23,488
And you will also provide us with that tracking number, as well.

576
00:34:29,195 --> 00:34:30,659
So, when the sample bottles arrive,

577
00:34:30,861 --> 00:34:35,912
we will first look at the 125-mL bottle

578
00:34:35,912 --> 00:34:41,467
to look at the sample under the scanning electron microscope to get the species identification.

579
00:34:42,477 --> 00:34:45,154
If it turns out to be a known toxin producer,

580
00:34:45,710 --> 00:34:48,639
we will then use the 1-Liter bottle for toxin testing.

581
00:34:49,497 --> 00:34:53,235
This is why it’s important NOT to add Lugol’s to the 1-Liter bottle,

582
00:34:53,235 --> 00:34:55,962
as that color change will interfere with the assay.

583
00:34:57,426 --> 00:35:00,911
The 30-mL bottle is used to get an actual cell count.

584
00:35:04,093 --> 00:35:08,184
Again, any bottles you are not asked to save for shipment will be reused.

585
00:35:09,042 --> 00:35:13,234
And remember to always enter data for every sample, 

586
00:35:13,739 --> 00:35:16,012
even if you do not see any targets.

587
00:35:24,194 --> 00:35:27,628
So, here is just a quick review of the protocols we have covered here today.

588
00:35:28,183 --> 00:35:31,113
If you have any further questions about the protocols,

589
00:35:31,416 --> 00:35:33,385
please don’t hesitate to email us.

590
00:35:36,567 --> 00:35:38,890
Now that you’ve completed Part 1 of the training,

591
00:35:38,890 --> 00:35:42,526
you must first complete a review of the information.

592
00:35:43,234 --> 00:35:45,708
This is done using a program called Menti.

593
00:35:46,466 --> 00:35:47,930
Your answers are anonymous, 

594
00:35:47,930 --> 00:35:52,930
but it gives us an idea of how well we are getting our information across to our volunteers. 

595
00:35:54,041 --> 00:35:57,021
Please email pmn@noaa.gov

596
00:35:57,021 --> 00:36:00,203
and request the special code to use for the review.

597
00:36:01,364 --> 00:36:03,486
Once you have completed that review quiz,

598
00:36:03,890 --> 00:36:06,920
you can then request to set up the second part of training.

599
00:36:07,879 --> 00:36:12,273
Part 2 will be done live via webinar, and according to your specific region.

600
00:36:13,081 --> 00:36:15,758
Possible dates will be determined on a rolling basis.

601
00:36:16,667 --> 00:36:18,182
When Part 2 is complete, 

602
00:36:18,586 --> 00:36:20,556
your supplies will then be shipped to you,

603
00:36:20,556 --> 00:36:23,132
and you can begin sampling for PMN.

604
00:36:25,051 --> 00:36:26,818
Again, if you have any questions,

605
00:36:26,818 --> 00:36:29,849
you can email us at pmn@noaa.gov

606
00:36:30,556 --> 00:36:33,283
or Jennifer.Maucher@noaa.gov

607
00:36:34,545 --> 00:36:35,051
Thank you!