Test Protocol for Legionella: Science Sunday

At our lab we are certified for two different methods for identifying and quantifying Legionella in samples. One is the general CDC method and the other is a method that is WAY more involved and only required by the State of New York.  Why the state of New York has to be extra special is beyond me, but we have been audited by them and found to do everything properly so we can do New York samples.

In the following post, I actually am doing both methods simultaneously because one client is from not New York, and one client is from New York. Many of the steps are similar. Here we go!

Here is nearly everything I need to test for both methods of Legionella. From left to right:  vacuum pump, 100ml sterilized tips, pipetter, erlenmeyer flask with rubber fitting, vortexer, sharpie, chlorine test strips, samples, skinny tubes, glass tubes, black agar plates, blue large tubes, Page's Saline, saline, NY acid, Legionella acid, cultures of Pseudomonas aeruginosa and Legionella pneumoniae, sterile water, 2um disposable filters.

Not pictured: 1 ml glass pipettes, pipette bulb, forceps, ethanol, lighter

First step, required for NY, optional for CDC.  Test the chlorine content of the water.

Good result! You can proceed with testing of the sample.

BAD RESULT.

Add sodium thiosulfate until result is good. In this case it was 100 ml of sample, so I added 100 ul at a time.

Fill large tubes with 10 ml of saline (Pages for NY, general for CDC)

Filter 100 ml of sample.

Dip the forceps into ethanol and SET THEM ON FIRE! (to sterilize) Once cool, use the forceps to transfer the filter paper to the blue eppendorf tubes.

VORTEX!  Just... generally for CDC, for 2 VERY LONG MINUTES for NY.

Yay!

Forceps and ethanol

For NY, you have to run positive and negative controls. Using the McFarland standards, you can guesstimate your concentration of bacteria in liquid by comparing the cloudiness.  Pseudomonas is the negative control, Legionella is duh, the positive.

To save time, vortex multiple tubes at once and cut vortexing time in half!

CDC:  Use bi plates with two similar agars on each side. Pipette 0.1 ml onto each side.

Spread that sample around. You can use the same spread from side 1 to 2, but not 2 to 1 and obviously not from one sample to the next.

NY samples go on a single type of agar (Side 2 but poured onto a whole plate). You plate 0.5 ml and spread.

Not pictured:  Each method has an acid treatment stage. This is pretty neat. The acid will kill amoebas that can hide Legionella. Yay! CDC has a 15 minute acid treatment.  NY has a 5 minute acid treatment in a different acid.

NY also has a 30 minute 50deg C heat treatment.

Each treatment also gets plated onto it's own plate.

In the end, CDC method uses 4 plates (regular and 3 acid) per sample, NY uses 3 plates per sample (regular, heat, acid) plus the controls.

All of those plates go into anaerobic jars.  You light the candle and tighten the lid, the candle burns all the oxygen. Those jars get incubated for 5 days (CDC) or up to 10 days (NY) at 35 deg C.

How we identify legionella coming up next!

BONUS SCIENCE

A picture of the product my company manufactures. It's a two sided dip stick. You dip it in a limited volume of water and bacteria or fungi stick to the agar. You can enumerate how much bacteria is in your volume of water.

Kid Pic Spam plus other stuff

An addendum to my previous post about Thinx, I'm also planning to try sea sponge tampons to see if they're more comfortable for riding (and pair with the thinx)  compared to my cup or disposable tampons.  The sponges are reusable for 3-6 months so you're still spending less and the product is somewhat renewable.

Anyway, I'll have the sponges in time for my next cycle, so I'll update then with how it all goes.


Hazel is making significant developmental progress. She can now easily crawl with her belly off the ground, is cruising around the house like a pro (when they walk but always hang on to stuff for balance) and has taken a single solo step on a few occassions.

She gets super  excited when she sees me pull out my carrier and loves being worn while I do stuff outside or go to the store. In general, she loves being the center of attention. Her favorite thing to say right now is Dada!  Which of course makes Eric very happy. I'm still getting her to work on Mama.

She loves Gwyn, Saffron and the cats and will screech excitedly/chase them.

She loves sitting in the high chair up above everyone. I think she likes watching them all.

Thinking about walking

We had a bout of sickness and she was diagnosed with RSV and had to have some breathing treatments. She's all better now though!

Sick baby passed out in the car. 

Using her toy to walk.

Two bottom teeth!

We did her 9 month photos a few days late.

Kaylee of course always wants to be in the pictures too.

Fun with colors at my job

The iridescent colors separate, it's really neat.

Kaylee insisted on riding in the snow.

Hazel is the center of attention while Kaylee is in gymnastics.

Kaylee is doing really well for her age group and she enjoys her once a week activity.

Eric is encumbered.

Side carred crib is also for cats!

A frequent sight.

Kisses for the kitty.

Sunrise on wednesday, my super busy day

I toured a school that I want Kaylee to attend for Kindergarten. It's a charter school that only accepts 40 students. Open enrollment ends Friday. This is the upper school (2nd through 8th grade)

Gym

Lower school (K-1)

Lower school recess. They are outside every day, regardless of weather.

Happy upside down baby.

Poking at Eric's moles.