What did MSO mean by this

[Naloac]

Bottom post of the previous page:

Completely ignoring the entire thing what the hell did he mean by the last line.



its a joke right? it has to be a joke

[Boot]

Hold up does the mra in your name actually mean men's rights activist, mso

Like card carrying mra

Why are you surprised? Isn't "men's right advocate" something that should describe all of us?

[MrStonedOne]

Hold up does the mra in your name actually mean men's rights activist, mso

Like card carrying mra

https://avoiceformen.com/feminist-gover ... h-culture/

Yes.

[EmpressMaia]

Hold up does the mra in your name actually mean men's rights activist, mso

Like card carrying mra

Why are you surprised? Isn't "men's right advocate" something that should describe all of us?

[Naloac]

[Ziiro]

no see you misunderstand, rules are used to punish others - not for the people who wrote them to be beholden to them

hope this helps

[Capsandi]

I post in /ss13g/ does that mean I'm retarded?

[Lacran]

I like to think I was quite a good headmin in my day, I trained alot of current admins or helped with their training. Was headmin successfully twice so I must have done something right :3 .Im sad that you've formed this opinion but what can I do

It's mainly because you arbitrarily gave rulings and were a spineless shit stirrer.

[Lacran]

It wouldn't be doxxing to say you mod subreddits on digg in a digg post. Though saying you sell children's toys would be, provided that info is not also from digg.

[sinfulbliss]

It wouldn't be doxxing to say you mod subreddits on digg in a digg post. Though saying you sell children's toys would be, provided that info is not also from digg.

whenever you post I think of that reddit guy with the fedora meme

[Lacran]

It wouldn't be doxxing to say you mod subreddits on digg in a digg post. Though saying you sell children's toys would be, provided that info is not also from digg.

whenever you post I think of that reddit guy with the fedora meme

You had me right up until you said you could think

[oranges]

I moderate r/ss13, what does that make me

[oranges]

I post in /ss13g/ does that mean I'm retarded?

yes

[Rohen_Tahir]

I moderate r/ss13, what does that make me

fat

[oranges]

I moderate r/ss13, what does that make me

fat

digg moderators don't get paid how are they going to eat enough to get fat

[dirk_mcblade]

Hold up does the mra in your name actually mean men's rights activist, mso

Like card carrying mra

Why are you surprised? Isn't "men's right advocate" something that should describe all of us?

Depends. Is there a specific issue or is MRA one of those "I should be assigned a girlfriend because I'm male" causes? Because while society in some matters is disadvantageous towards men, men also straight up commit like 98% of the murders, so which side is more in the wrong?

[dirk_mcblade]

I moderate r/ss13, what does that make me

fat

digg moderators don't get paid how are they going to eat enough to get fat

Poor people are more likely to be obese in America. I don't know what percentage of income goes towards food in the shire but in the USA you get food stamps if you've got no income or savings.

[Boot]

Hold up does the mra in your name actually mean men's rights activist, mso

Like card carrying mra

Why are you surprised? Isn't "men's right advocate" something that should describe all of us?

Depends. Is there a specific issue or is MRA one of those "I should be assigned a girlfriend because I'm male" causes? Because while society in some matters is disadvantageous towards men, men also straight up commit like 98% of the murders, so which side is more in the wrong?

I think more broadly. MRA being shorthand for "men's right advocate" just seems more like a descriptor. Like do you think that men shouldn't have to be the military property of the state to die for at the drop of a hat? Would you advocate for that position? Then it would be accurate to describe you as a MRA. Do you think that men should be allowed to keep all of their skin? MRA.

[sinfulbliss]

Hold up does the mra in your name actually mean men's rights activist, mso

Like card carrying mra

Why are you surprised? Isn't "men's right advocate" something that should describe all of us?

Depends. Is there a specific issue or is MRA one of those "I should be assigned a girlfriend because I'm male" causes? Because while society in some matters is disadvantageous towards men, men also straight up commit like 98% of the murders, so which side is more in the wrong?

I’m just guessing but I reckon MRA was a response to the more radical wing of WRA that sorta started to go more into the “men are the problem” territory.

Also I think men committing all the murders isn’t exactly a counterpoint to the “men are disadvantaged in society” argument. Not that I think they are, but why would that be an advantage!!

Men commit most of the murders because of testosterone and a higher proclivity towards violence as a result, pretty sure. Probably also partially a cultural thing with the glorification of violence as a form of solving issues/problems as a man.

[dirk_mcblade]

Hold up does the mra in your name actually mean men's rights activist, mso

Like card carrying mra

Why are you surprised? Isn't "men's right advocate" something that should describe all of us?

Depends. Is there a specific issue or is MRA one of those "I should be assigned a girlfriend because I'm male" causes? Because while society in some matters is disadvantageous towards men, men also straight up commit like 98% of the murders, so which side is more in the wrong?

I think more broadly. MRA being shorthand for "men's right advocate" just seems more like a descriptor. Like do you think that men shouldn't have to be the military property of the state to die for at the drop of a hat? Would you advocate for that position? Then it would be accurate to describe you as a MRA. Do you think that men should be allowed to keep all of their skin? MRA.

Well I mean if murders are overwhelmingly committed by men then they're simply much more suitable for killing others and therefore exactly the kind of people you want in a combat unit. How would a draft involving women work? Would they go into noncombat roles to free up more men for fighting? Or would they be integrated into the front lines? The latter would be retarded based on common sense and the marine corps study on this.

Hold up does the mra in your name actually mean men's rights activist, mso

Like card carrying mra

Why are you surprised? Isn't "men's right advocate" something that should describe all of us?

Depends. Is there a specific issue or is MRA one of those "I should be assigned a girlfriend because I'm male" causes? Because while society in some matters is disadvantageous towards men, men also straight up commit like 98% of the murders, so which side is more in the wrong?

I’m just guessing but I reckon MRA was a response to the more radical wing of WRA that sorta started to go more into the “men are the problem” territory.

Also I think men committing all the murders isn’t exactly a counterpoint to the “men are disadvantaged in society” argument. Not that I think they are, but why would that be an advantage!!

Men commit most of the murders because of testosterone and a higher proclivity towards violence as a result, pretty sure. Probably also partially a cultural thing with the glorification of violence as a form of solving issues/problems as a man.

Men commit the vast majority of murders globally, it's not a cultural issue. If the percentage of men being victims of murder drops below 98% then the victims are disproportionately female, which means women are murdered in excess of themselves murdering others. In some places, Hong Kong and South Korea for example, women are more likely to be murdered than men. I think that's fucked up. I don't think being a murderer is an advantage in society, but I think given how the vast majority of criminality is done by men it'd be rational to scrutinize the activities of men. It's also hard to say the men are disadvantaged moreso than women regarding murder if women are more often murdered by men than the other way around.
Like, I don't consider myself a feminist, but some of the aspects of masculinity is pretty indefensible while nonetheless existing as basically a force of nature.

[dendydoom]

firstly, this thread sucks.

secondly, these discussions always hide some inner political belief that people are subtly deferring to.

as usual i will use my personal life experience to tell a story about this:

i'm a woman and when i was younger i spent a lot of time online and wanted to be liked by my peers and accepted into "the culture" of 2008 era 4chan shitposting. i would tell intolerant jokes, say bigoted things and mock "normies" because they were the outside and i needed social cred to elevate myself on the inside.

years later in my late teen years i started hanging out with other women who were politically active and they taught me about feminism and i had a phase where i was one of those "men are the problem" types and i was just as much of an insufferable jackass but now i hung out on the other side of the divide.

then a few years later, in my early-mid 20s, i had calmed down because i had made friends with men, women, normies and internet dwellers all alike. i started to find it difficult to tell those same jokes, to hold those same beliefs against men or women because every time i did i would think of my friends, their life experiences, the problems they faced and i knew it would hurt them to hear me say those things and it became impossible to be so reductive to the true breadth and depth of their struggles and experiences. i care about them, no matter who they are, and i want to help them self actualize and be free from the things causing them pain.

what i'm ultimately trying to say is that it does no side any good to paint the other with a broad brush. i still believe in intersectionality: the idea that all people are a big ball of different types of privilege and discrimination based on a wide array of attributes. men wield the majority of power in society, but that power does not benefit all men. access to mental healthcare is worse for men, suicide affects them more, it is harder for them to show emotional vulnerability, they are the ones who are made to take dangerous jobs, they are the ones that die in wars, they are the ones who often lose access to their children in divorce, the list goes on and on. obviously it goes without saying that there is a whole host of issues that affect women too. it doesn't mean any of that stops existing like it's some sort of competition over who gets fucked over more. we all do. the thing that will change you and better you in this regard is to make friends with everyone, even those you might disagree with. they will see you as a real person, not some political opposition that's dangerous to them, and you will see the same in them too, and both of you will grow from sharing your experiences and your struggles.

[sinfulbliss]

Well I mean if murders are overwhelmingly committed by men then they're simply much more suitable for killing others and therefore exactly the kind of people you want in a combat unit. How would a draft involving women work? Would they go into noncombat roles to free up more men for fighting? Or would they be integrated into the front lines? The latter would be retarded based on common sense and the marine corps study on this.

I genuinely don't see the connection between men committing murders and the claim they are advantaged... because they would be wanted for a combat unit? That sucks who would want to be in a combat unit!
And yeah obviously drafts should be preferential to men, sucks but that's how it has to be so long as wars exist.

Men commit the vast majority of murders globally, it's not a cultural issue. If the percentage of men being victims of murder drops below 98% then the victims are disproportionately female, which means women are murdered in excess of themselves murdering others. In some places, Hong Kong and South Korea for example, women are more likely to be murdered than men. I think that's fucked up. I don't think being a murderer is an advantage in society, but I think given how the vast majority of criminality is done by men it'd be rational to scrutinize the activities of men. It's also hard to say the men are disadvantaged moreso than women regarding murder if women are more often murdered by men than the other way around.
Like, I don't consider myself a feminist, but some of the aspects of masculinity is pretty indefensible while nonetheless existing as basically a force of nature.

This is just an incredibly bizarre, tangled argument you've constructed. First of all men commit murder more =/= Women get murdered more. The murder victim rate is like, overwhelmingly male, around 5:1 in the US. That's kind of a nothing-statement though, obviously men are killed more because they're more violent and prone to violent crime than women on average. But yeah have no idea why you'd make this assumption.

I'll concede it's probably more of a biological issue than a cultural one, considering the people who commit violent crimes correlate with the people who are the most aggressive and antisocial, which is pretty much all men. That's probably due to a mix of biology and culture though.

it does no side any good to paint the other with a broad brush

Basically this

[yobihodazine2]

Men are simply more capable of, and susceptible to, acting on their violent urges. Women are more likely to solve their problems in other ways. There are exceptions. If women were as physically capable as men I have no doubt their murder rate would even out. I mean, some women aren't afraid to take any advantage given to them.

[dirk_mcblade]

I genuinely don't see the connection between men committing murders and the claim they are advantaged... because they would be wanted for a combat unit? That sucks who would want to be in a combat unit!

This is just an incredibly bizarre, tangled argument you've constructed. First of all men commit murder more =/= Women get murdered more. The murder victim rate is like, overwhelmingly male, around 5:1 in the US. That's kind of a nothing-statement though, obviously men are killed more because they're more violent and prone to violent crime than women on average. But yeah have no idea why you'd make this assumption.

Loads of people already volunteer for combat units so it apparently contains some value to someone. Navy SEALS aren't conscripts for example. But at any rate you should ask the people pushing for women to be in combat units, not me. Shit like https://www.brookings.edu/essay/women-w ... ed-forces/ frames it as an accessibility problem for women in the military and refer to it as a "milestone" that they began to be allowed to do so recently. So don't ask me, ask the people pushing for it. It's apparently treated like it's an advantage. I think it's batshit crazy but I'm not the guy being a proponent of it out of "fairness". My perspective is if you're going to issue a draft you want to assign the most effective people to the direct combat roles and it's not an argument I base on equitability but because the survival of the given country would be at stake. Or at least it should be. The most recent drafts in the USA and Russia concerned force projection rather than responses to being attacked first.

Women get killed by men far more often than men get killed by women, therefore it's not surprising that at least some women regard men as the problem. Also the statement of "obviously men are killed more because they're more violent and prone to violent crime than women on average" is exactly my point. If women are murdered 20% of the time but are murderers only 2% of the time that implies women are being targeted disproportionately despite being less violent than men.

[sinfulbliss]

I genuinely don't see the connection between men committing murders and the claim they are advantaged... because they would be wanted for a combat unit? That sucks who would want to be in a combat unit!

This is just an incredibly bizarre, tangled argument you've constructed. First of all men commit murder more =/= Women get murdered more. The murder victim rate is like, overwhelmingly male, around 5:1 in the US. That's kind of a nothing-statement though, obviously men are killed more because they're more violent and prone to violent crime than women on average. But yeah have no idea why you'd make this assumption.

Loads of people already volunteer for combat units so it apparently contains some value to someone. Navy SEALS aren't conscripts for example. But at any rate you should ask the people pushing for women to be in combat units, not me. Shit like https://www.brookings.edu/essay/women-w ... ed-forces/ frames it as an accessibility problem for women in the military and refer to it as a "milestone" that they began to be allowed to do so recently. So don't ask me, ask the people pushing for it. It's apparently treated like it's an advantage. I think it's batshit crazy but I'm not the guy being a proponent of it out of "fairness". My perspective is if you're going to issue a draft you want to assign the most effective people to the direct combat roles and it's not an argument I base on equitability because the survival of the given country would be at stake.

Women get killed by men far more often than men get killed by women, therefore it's not surprising that at least some women regard men as the problem. Also the statement of "obviously men are killed more because they're more violent and prone to violent crime than women on average" is exactly my point. If women are murdered 20% of the time but are murderers only 2% of the time that implies women are being targeted disproportionately despite being less violent than men.

That’s like saying it’s primarily Muslims who commit terrorist attacks on the US therefore it makes sense to regard Muslims as the problem.

No man you can’t broad brush an entire group of people over the actions of the most extreme portion of them, that’s bad news. Your statistics are also completely out of whack, that’s not how proportions work. If women are murdered 20% of the time but only murderers 2% of the time then that means two things: mostly men are killed, and mostly men commit murder. It doesn’t mean women are disproportionately murdered, it means there are more women killed by murder than women who commit murder themselves.

[dirk_mcblade]

I genuinely don't see the connection between men committing murders and the claim they are advantaged... because they would be wanted for a combat unit? That sucks who would want to be in a combat unit!

This is just an incredibly bizarre, tangled argument you've constructed. First of all men commit murder more =/= Women get murdered more. The murder victim rate is like, overwhelmingly male, around 5:1 in the US. That's kind of a nothing-statement though, obviously men are killed more because they're more violent and prone to violent crime than women on average. But yeah have no idea why you'd make this assumption.

Loads of people already volunteer for combat units so it apparently contains some value to someone. Navy SEALS aren't conscripts for example. But at any rate you should ask the people pushing for women to be in combat units, not me. Shit like https://www.brookings.edu/essay/women-w ... ed-forces/ frames it as an accessibility problem for women in the military and refer to it as a "milestone" that they began to be allowed to do so recently. So don't ask me, ask the people pushing for it. It's apparently treated like it's an advantage. I think it's batshit crazy but I'm not the guy being a proponent of it out of "fairness". My perspective is if you're going to issue a draft you want to assign the most effective people to the direct combat roles and it's not an argument I base on equitability because the survival of the given country would be at stake.

Women get killed by men far more often than men get killed by women, therefore it's not surprising that at least some women regard men as the problem. Also the statement of "obviously men are killed more because they're more violent and prone to violent crime than women on average" is exactly my point. If women are murdered 20% of the time but are murderers only 2% of the time that implies women are being targeted disproportionately despite being less violent than men.

That’s like saying it’s primarily Muslims who commit terrorist attacks on the US therefore it makes sense to regard Muslims as the problem.

No man you can’t broad brush an entire group of people over the actions of the most extreme portion of them, that’s bad news. Your statistics are also completely out of whack, that’s not how proportions work. If women are murdered 20% of the time but only murderers 2% of the time then that means two things: mostly men are killed, and mostly men commit murder. It doesn’t mean women are disproportionately murdered, it means there are more women killed by murder than women who commit murder themselves.

"It's not understandable that some people would regard men as the problem because we kill each other more often than the sex that's generally non-combatative (but we still kill a lot of them too)."

[yobihodazine2]

It's absolutely understandable. Which is why you need to guard your heart, you never know what kind of assumptions someone might bring into your relationship. By the way this girl I was seeing once said to me "you seem like the kind of person I'd want to be murdered by."

[BlueMemesauce]

In some places, Hong Kong and South Korea for example, women are more likely to be murdered than men. I think that's fucked up.

Hong Kong has a female homicide rate of 52.9% while South Korea has a rate 52.5%. Yes women are technically more likely to be murdered, but it's only 2%, it's practically equal. It's nothing compared to the vastly higher percent of male homicides in countries like the United States.
You're implying Hongkongers and Korean people just hate women way more than in the United States, but that's not the case. It's actually because Hong Kong and South Korea have the lowest homicide rates in the world. The rates being equal is mostly because less male gang members are killing men (Although I don't have any evidence for that, it's just my assumption), which is probably why the rates of homicide by gender are equal. It's not fucked up at all, it's a good thing when they're equal because that means neither of them are disproportionately killed. It seems you just value women more than men, which is why you think it's fucked up when women are killed at a rate equal to men. Edit: Okay I preemptively apoligize for saying that you're sexist if you actually meant that women were being killed at a way higher rate is wrong, rather than those specific examples of Hong Kong and South Korea. But what I mean is that since society pressures us to value women more than men, it may influence us to see a relatively low amount of women being disproportionately killed as worse than a high amount of men.

[dirk_mcblade]

In some places, Hong Kong and South Korea for example, women are more likely to be murdered than men. I think that's fucked up.

Hong Kong has a female homicide rate of 52.9% while South Korea has a rate 52.5%. Yes women are technically more likely to be murdered, but it's only 2%, it's practically equal. It's nothing compared to the vastly higher percent of male homicides in countries like the United States.
You're implying Hongkongers and Korean people just hate women way more than in the United States, but that's not the case. It's actually because Hong Kong and South Korea have the lowest homicide rates in the world. The rates being equal is mostly because less male gang members are killing men (Although I don't have any evidence for that, it's just my assumption), which is probably why the rates of homicide by gender are equal. It's not fucked up at all, it's a good thing when they're equal because that means neither of them are disproportionately killed. It seems you just value women more than men, which is why you think it's fucked up when women are killed at a rate equal to men. edit: okay I apologize that it might be wrong of me to say that you're sexist if you actually saw women were being killed at a way higher rate.

It's generally not women killing men in most countries (perhaps all even but I won't bother going that far because I'd have to confirm there isn't even one country like that). I don't see how it's sexist to point out that it's more common for men to kill women than the other way around.

[BlueMemesauce]

I agree with you on that point, that wasn't the part of your argument I disagreed with. You can see I only quoted the sentence where you said it was fucked up more women were being killed in Hong Kong and South Korea.

Edit: I also apologize for my bad habit of editing posts after sending them

[sinfulbliss]

"It's not understandable that some people would regard men as the problem because we kill each other more often than the sex that's generally non-combatative (but we still kill a lot of them too)."

Correct! It isn’t understandable to regard an entire class of people as a “problem.” Not men, not women, not black people, not Muslim people, not gay people, etc. I’m sure if I took any one of those groups and called them a “problem” for whatever disproportionate bad thing I wanted to find, you’d find it incredibly awful of me to say.

[oranges]

they've got allen wrenches gerbil feeders toilet seats electric heaters trash compactors juice extractor shower rods and water meters walkie-talkies copper wires safety goggles radial tires bb pellets rubber mallets fans and dehumidifiers picture hangers paper cutters waffle irons window shutters paint removers window louvres masking tape and plastic gutters kitchen faucets folding tables weather stripping jumper cables hooks and tackle grout and spackle power foggers spoons and ladles pesticides for fumigation high-performance lubrication metal roofing water proofing multi-purpose insulation air compressors brass connectors wrecking chisels smoke detectors tire gauges hamster cages thermostats and bug deflectors trailer hitch demagnetizers automatic circumcisers tennis rackets angle brackets duracells and denergizers soffit panels circuit breakers vacuum cleaners coffee makers calculators generators matching salt and pepper shakersthey've got allen wrenches gerbil feeders toilet seats electric heaters trash compactors juice extractor shower rods and water meters walkie-talkies copper wires safety goggles radial tires bb pellets rubber mallets fans and dehumidifiers picture hangers paper cutters waffle irons window shutters paint removers window louvres masking tape and plastic gutters kitchen faucets folding tables weather stripping jumper cables hooks and tackle grout and spackle power foggers spoons and ladles pesticides for fumigation high-performance lubrication metal roofing water proofing multi-purpose insulation air compressors brass connectors wrecking chisels smoke detectors tire gauges hamster cages thermostats and bug deflectors trailer hitch demagnetizers automatic circumcisers tennis rackets angle brackets duracells and denergizers soffit panels circuit breakers vacuum cleaners coffee makers calculators generators matching salt and pepper shakersthey've got allen wrenches gerbil feeders toilet seats electric heaters trash compactors juice extractor shower rods and water meters walkie-talkies copper wires safety goggles radial tires bb pellets rubber mallets fans and dehumidifiers picture hangers paper cutters waffle irons window shutters paint removers window louvres masking tape and plastic gutters kitchen faucets folding tables weather stripping jumper cables hooks and tackle grout and spackle power foggers spoons and ladles pesticides for fumigation high-performance lubrication metal roofing water proofing multi-purpose insulation air compressors brass connectors wrecking chisels smoke detectors tire gauges hamster cages thermostats and bug deflectors trailer hitch demagnetizers automatic circumcisers tennis rackets angle brackets duracells and denergizers soffit panels circuit breakers vacuum cleaners coffee makers calculators generators matching salt and pepper shakersthey've got allen wrenches gerbil feeders toilet seats electric heaters trash compactors juice extractor shower rods and water meters walkie-talkies copper wires safety goggles radial tires bb pellets rubber mallets fans and dehumidifiers picture hangers paper cutters waffle irons window shutters paint removers window louvres masking tape and plastic gutters kitchen faucets folding tables weather stripping jumper cables hooks and tackle grout and spackle power foggers spoons and ladles pesticides for fumigation high-performance lubrication metal roofing water proofing multi-purpose insulation air compressors brass connectors wrecking chisels smoke detectors tire gauges hamster cages thermostats and bug deflectors trailer hitch demagnetizers automatic circumcisers tennis rackets angle brackets duracells and denergizers soffit panels circuit breakers vacuum cleaners coffee makers calculators generators matching salt and pepper shakersthey've got allen wrenches gerbil feeders toilet seats electric heaters trash compactors juice extractor shower rods and water meters walkie-talkies copper wires safety goggles radial tires bb pellets rubber mallets fans and dehumidifiers picture hangers paper cutters waffle irons window shutters paint removers window louvres masking tape and plastic gutters kitchen faucets folding tables weather stripping jumper cables hooks and tackle grout and spackle power foggers spoons and ladles pesticides for fumigation high-performance lubrication metal roofing water proofing multi-purpose insulation air compressors brass connectors wrecking chisels smoke detectors tire gauges hamster cages thermostats and bug deflectors trailer hitch demagnetizers automatic circumcisers tennis rackets angle brackets duracells and denergizers soffit panels circuit breakers vacuum cleaners coffee makers calculators generators matching salt and pepper shakers

[Lacran]

The mens health takes here are so fucking bad stop holy shit

You can't use biological determinism for crime statistics if violent crime statistics vary wildly between countries. You're also ignoring how serious male violence is treated compared to female violence and the chance they'd actually be prosecuted.

Crime is overwhelmingly socio-economic. There are some biological factors sure but how we view those is more statistically relevant.

[yobihodazine2]

I needed that oranges

[carshalash]

Wait, swept has only played 46 rounds?

[sinfulbliss]

You can't use biological determinism for crime statistics if violent crime statistics vary wildly between countries.

No one mentioned biological determinism except you, there’s a difference between “biology makes a difference in your proclivity to violence” and “biology destines people to be violent.” Also male biology doesn’t really differ between countries so again why is it predominantly male?

Crime is overwhelmingly socio-economic. There are some biological factors sure but how we view those is more statistically relevant.

Except this isn’t a discussion about crime statistics in general it’s a discussion about gendered crime difference so that’s why it’s what’s being talked about.

[Lacran]

You can't use biological determinism for crime statistics if violent crime statistics vary wildly between countries.

No one mentioned biological determinism except you, there’s a difference between “biology makes a difference in your proclivity to violence” and “biology destines people to be violent.” Also male biology doesn’t really differ between countries so again why is it predominantly male?

Crime is overwhelmingly socio-economic. There are some biological factors sure but how we view those is more statistically relevant.

Except this isn’t a discussion about crime statistics in general it’s a discussion about gendered crime difference so that’s why it’s what’s being talked about.

The vast Majority of men aren't violent criminals, so you need to look at the crime as a whole. Why is someone becoming violent?

Crime statistics is relevant when you are talking about crime frequency between groups of people. If you say the reasons there is a statistical difference is because one group has more testosterone and is physically larger, you are saying biology is the determiner of the difference. Its like how lesbian couples have a higher domestic violence rate. Is it just that lesbians are more likely, or is the way we view and treat these situations fundamentally different?

Men being larger and having more testosterone is definitely a factor. But our mindsets around these biological realities are a lot more impactful than the biology themselves. A small woman assaulting a larger man is viewed differently and is far less likely to result in an assault charge.

The reason I bring up crime in general and why it is different between countries is because males are not that different between countries, so you can't just look at how men are different to women, you have to look at how different environments affect these groups and their likelihood to do crime and how that society views that. Thailand's female incarceration rate is 20% whereas pakistan's is 1% so would the arguement be that men and women are somehow fundamentally different there?

[Not-Dorsidarf]

hey guys im back from holiday howve you been keeping the forum


oh no

[conrad]

This thread is rancid.

[Rohen_Tahir]

The wasted chewing gum bacteriome
Leila Satari,1 Alba Guillén,1 Àngela Vidal-Verdú,1 and Manuel Porcarcorresponding author1,2

Abstract

Here we show the bacteriome of wasted chewing gums from five different countries and the microbial successions on wasted gums during three months of outdoors exposure. In addition, a collection of bacterial strains from wasted gums was set, and the biodegradation capability of different gum ingredients by the isolates was tested. Our results reveal that the oral microbiota present in gums after being chewed, characterised by the presence of species such as Streptococcus spp. or Corynebacterium spp., evolves in a few weeks to an environmental bacteriome characterised by the presence of Acinetobacter spp., Sphingomonas spp. and Pseudomonas spp. Wasted chewing gums collected worldwide contain a typical sub-aerial biofilm bacteriome, characterised by species such as Sphingomonas spp., Kocuria spp., Deinococcus spp. and Blastococcus spp. Our findings have implications for a wide range of disciplines, including forensics, contagious disease control, or bioremediation of wasted chewing gum residues.
Subject terms: Environmental microbiology, Applied microbiology
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Introduction

Chewing gums may have been used for thousands of years, since wood tar from the Mesolithic and Neolithic periods have been found with tooth impressions, which suggests a role in teeth cleaning as well as its usage as early adhesives1, 2. The first modern chewing gum was introduced in the market in the late 19th3 and chewing gums are today vastly consumed worldwide: it is estimated that Iran and Saudi Arabia are the countries with the highest chewing gum consumption, where 80% of the population are regular chewing gum consumers4. Moreover, global online surveys on gum intake conducted in Europe5 and United States6 displayed similar chewing gum patterns among them, where more than 60% of adolescents and adults had chewed gums in the last 6 months before the survey and the mean intakes ranged from 1 to 4 pieces of chewing gum per day. Significantly, Hearty et al. (2014)5 reported the lowest chewing gum intake (46%) in the United Kingdom. Finally, the value of chewing gum trade has been estimated as more than 30 billion U.S. dollars in 20197.

Chewing gums are generally composed of two phases: the water-insoluble phase (gum base) and the water-soluble phase, which can be made of sugar (sugar chewing gums) or sugar alcohols such as polyols (sugar-free chewing gums). Some chewing gums present a solid coat, which is involved in flavour release as well as in chewing gum protection to physicochemical agents, that can be specified as a third phase8. The main component of any chewing gum is the gum base (20–30%), that is not edible, nor digestible, but allows chewing, during which added flavours and sweeteners are released. Indeed, chewing gum can be chewed for a long period without any structural modifications because of the water-insoluble property of the gum base9. Gum base can be produced from either natural polymers, such as latex or waxes, or synthetic polymers, particularly polyvinyl acetate (15–45%)—a key ingredient in chewing gum formulation—and synthetic elastomers (10–30%) including co-polymers of butadiene-styrene, isobutylene-isoprene as well as, polyethylene, polyisobutylene and polyisoprene8. Hence, this inert part of the formula constitutes the support for the water-soluble components which consist of: (i) sweeteners, whether sugar or sugar alcohols that constitute the 60% of the chewing gum; (ii) humectants, such as glycerine; (iii) antioxidants, supplemented to avoid oxidation of other ingredients; (iv) colours, flavours and organic acids, added to define an specific taste of the chewing gum; and (v) optionally, “active” ingredients such as nicotine in chewing gums as an alternative to smoking8, 9. The particular amounts of these components in the chewing gum formula are a well-kept secret of each confectionery industry. As hinted before, sweeteners comprise more than half of the chewing gum composition. Sucrose, dextrose and glucose syrup are the most frequently used in sugar-containing gums. However, most of chewing gums present in the European market are sweetened with polyols (sugar alcohols) such as xylitol, sorbitol, mannitol, maltitol and isomalt10, 11 as well as artificial sweeteners such as aspartame12, all of which being labelled as sugar-free chewing gums. The effect of the sugar-free chewing gums in the control of dental disease, salivary pH, and the oral microbiome has been reported10, 13–15.

Wasted chewing gums are often improperly discarded and end up as long-lasting residues on both indoor and outdoor pavements and surfaces. Local councils spend millions of euros cleaning up gum residues from the pavement. For instance, it is estimated that in the UK the annual cost of cleaning up wasted gums form streets is almost 70 million euros16. Besides, there is a big concern regarding chewing gum residues stuck to historic buildings or art works, which contribute negatively to its conservation17. Moreover, the popularity of chewing gums as well as the widespread presence of those long-lasting residues have allowed using wasted chewed gums for human genetic analysis in criminology18 and archaeology19, 20. It has to be stressed, though, that beyond its contents in the consumer’s DNA, used chewed gum can contain an important fraction of the oral microbiome21, toxins22, and some opportunistic pathogens such as Streptococcus spp. and Actinomyces spp.23, 24, which remain trapped in the sticky residue, and whose survival over time has received only limited attention to date17, 21. Wasted chewing gums are considered as environmental pollutants, mainly for aesthetic reasons, and their removal from pavements can be economically expensive, and time-consuming25. To date, most studies aiming at improving wasted chewing gums cleaning have mainly focused on the production of less adhesive, water-soluble and degradable chewing gums22, 26.

The present work describes a complete characterization on the bacterial contents of wasted chewed gum, by using culture-dependent and -independent techniques. We have studied the microbial content of wasted chewing gums sampled in different locations worldwide as well as the distribution of bacteria depending on the depth (surface, intermediate and bottom layers of the residue) and conducted a dynamic study to shed light on the microbial succession that takes place in the chewing gum during the first weeks after its disposal on an outdoor surface. On the other hand, we have screened the biodegradation capability of the gum ingredients of a collection of bacterial strains we isolated from chewing gum residues. Our results have implications in fields such as criminology, contagious disease control, waste management and bioremediation.
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Results
Bacterial communities in wasted chewing gums analysed through NGS

The bacterial diversity of eight chewing gum samples collected in five different countries was analysed through NGS as described in the Materials and Methods section and the taxonomic profiles are shown in Fig. 1. Two chewing gum samples collected in Spain, France, and Singapore were analysed; while from Greece and Turkey one chewing gum sample was analysed. The bacterial profile deduced by the analysis of the pool of 16S rRNA genes reveals relatively similar bacterial profiles, but yet with differences in some genera. Interestingly, one of the samples from Singapore displayed a very high biodiversity with a total of 427 identified taxa, which results in a low relative abundance of the most common genera, being approximately the 15% in this sample. However, the number of samples per country being too small to draw conclusions at the geographic level, these results have to be considered as a first approach on the chewing gum bacteriome. Chewing gum samples from France (Paris) and Turkey (Istanbul) proved also rather diverse representing the most common genera the 49% and 42% of total abundance, respectively.
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Figure 1

The taxonomic profiles of wasted chewing gum samples collected from outdoor locations in five countries. The most abundant taxa from each location are shown in pie charts. Figure created with Adobe Illustrator CS6 (www.adobe.com).

Although the eight samples exhibited an important variation among locations, several genera were found in all the analysed samples (Kocuria, Sphingomonas, Deinococcus, Blastococcus, Skermanella, Hymenobacter, Modestobacter, Roseomonas, Rubellimicrobium, Methylobacterium, Uncultured Sphingomonas, Paracoccus and Nocardiodes). Their relative abundance changed significantly among samples. The genus Kocuria with a 54.18% frequency and Sphingomonas with a 38.9%, were the most abundant genera in one of the samples from Singapore and France, whereas in other samples, such as Greece, the most frequent genera were Deinococcus with a 25.2% frequency, followed by Kocuria, Pseudokineococcus and Modestobacter with relative abundances from 10 to 16%. In samples from Valencia, other frequent genera were Blastococcus, Nesterenkonia and Hymenobacter with a 23.2%, 27.8% and 14.4% relative abundance, respectively. Finally, the genus Skermanella was the most abundant one in the chewing gum from Turkey (20.9%). Arthrobacter spp. was identified in all samples except the two samples from Singapore. Similarly, Pseudomonas spp. was found in all samples except in the one from Greece. Interestingly, in the two Mediterranean locations, three genera constituted approximately 25% of the biodiversity. These genera were: Blastococcus and Nesterenkonia from Valencia, Spain; and Deinococcus from Spetses Island, Greece. The presence of genus Nesterenkonia was found exclusively in the two samples from Spain. In addition, the genus Curtobacterium was found mainly in Turkey and in a lower abundance in France (Paris).

In one case, a single gum sample was collected and divided in situ in three different fractions that were processed independently. The sample, located in one of the outdoor car parks of the Scientific Park of the University of Valencia, was collected in three consecutive sections, corresponding to the upper, medium and bottom layers of the residue. As shown in Fig. 2, the taxonomic profiles of the three sub-samples proved almost identical in composition. The surface layer, which is the most exposed to environmental conditions, presented a higher abundance of chloroplasts. The main genus identified in the three samples was Curtobacterium with a relative abundance between 32–47%, being more frequent in the intermediate layer. Other, less frequent genera were Sphingomonas, which was the second most abundant genus and represented more than 16% of the biodiversity; Hymenobacter with an 8%; and Kineococcus, with a 4–6%. Finally, Massilia, an uncultured Sphingomonadaceae, Methylobacterium, Aureimonas and members of the Rhizobium clade (Allorhizobium, Neorhizobium, Pararhizobium and Rhizobium) had similar frequencies, of 1.5–5%. The remaining genera presented similar abundances and constituted approximately 9% of the total biodiversity in each layer.
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Figure 2

Comparison between the bacterial communities of three layers from a chewed gum residue. (a) Upper layer, (b) medium layer, and (c) bottom layer (described in M&M) showing small variation in biodiversity.
Colonisation process of wasted chewing gums

A specific experiment was carried out to shed light on the microbial successions taking place in wasted chewing gum once discarded. Thirteen gums were chewed and placed on an outdoor pavement over a period of up to 12 weeks and high-throughput 16S rRNA sequencing was carried out to follow the dynamic changes in their bacterial contents. The shape of rarefaction curves at OTU level revealed a good coverage of the actual bacterial diversity (Supplementary Fig. 1). The high throughput sequencing and analysis of the 16S rRNA gene amplicons from all the samples revealed shifts in bacterial diversity in time (Fig. 3a), reaching the highest alpha diversity values after 6–8 weeks. The most abundant genera (Fig. 3b) included the genus Streptococcus, with a relative abundance of more than 25% in the control sample (which had its total DNA immediately extracted after being chewed). The relative frequency of this genus slowly decreased in time and reached its lowest abundance by the ninth week. Other frequent genera found in the control sample were the oral microbiome members Rothia, Haemophilus, Corynebacterium, Veillonella, Actinomyces and, to a lesser extent, Granulicatella and Gemella. All these genera remained detectable in the analysed samples throughout the whole experiment, but they clearly decreased in time, although with different temporal patterns. Interestingly, the relative abundance of non-oral, environmental genera, such as Rubellimicrobium, Sphingomonas, Acinetobacter, and Pseudomonas increased with the time the gums were kept outdoors, reaching maximum values of 12%, 19%, 23% and 16% respectively, after sixth, eighth, eleventh, and twelfth weeks each.
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Figure 3

Dynamic experiment showing the variation of the chewed gum microbial communities in a twelve weeks period. (a) Alpha diversity shows the richness of the samples based on the Richness, Shannon, and Simpson index, (b) Clustered-Bar chart represents the modification of the oral bacterial profile over time, according to 16S rRNA gene monitoring. (c) Beta diversity (PCoA) based on Bray–Curtis illustrates correlations among the bacterial genera in different samples. Distances to the linear statistical correlation and the colours indicate the similarity of the microbial communities affected by environmental factors or removing time during the three-month period of the experiment.

A Principal Coordinate Analysis (PCoA) proved that the bacterial communities in wasted chewing gums during a period of 12 weeks clustered in three groups (Fig. 3c). These groups corresponded to samples collected after 1–4; 5–7; and 9–12 weeks, thus indicating a clear association between taxonomic profile similarities and time of outdoor exposition.

In order to identify the fate of the most common genera in the control sample in time, their relative abundance as well as that of the most frequent genera at the end (12th week) of the experiment were compared, as shown in Fig. 4. The four most abundant genera in the control sample were Streptococcus, Corynebacterium, Haemophilus, and Rothia, which constituted 56% of biodiversity of the sample (Fig. 4a). On the other hand, sample 12 included 9 main genera (51% of the reads) corresponding to environmental bacteria. In fact, and as Fig. 4 shows, the presence of the most abundant taxa in the control sample decreased significantly and was substituted by a soil-related profile in which the most frequent genera were Acinetobacter, Sphingomonas and Pseudomonas. The ecological succession of seven selected environmental genera (Sphingomonas, Rubellimicrobium, Craurococcus, Granulicatella, Deinococcus, Hymenobacter, and Kocuria) were monitored over time (Supplementary Fig. 2). All these genera, except Granulicatella, with a 4% abundance at the start, were not present in the control sample or only in a very low quantity, as the case of Sphingomonas and Rubellimicrobium, which showed an upward trend over time, peaking in the intermediate stages and stabilizing towards the end of the monitoring.
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Figure 4

Comparison between the most frequent genera in (a) a control gum directly analysed after being chewed and (b) a sample taken after 12 weeks placed outdoors.
Culture-dependent experiments

Strain collection from wasted chewing gums

Different chewing gum samples from the vicinity of our laboratory (Valencia, Spain) were ground and spread on different microbiological media. A total of 21 bacterial colonies, were selected, isolated and identified by 16S rRNA gene sequencing. The isolates selected from aerobically-incubated Petri dishes belonged to the following genera; Curtobacterium (S1-1), Pantoea (S1-2), Microbacterium (S1-3 and S1-6), Pseudomonas (S1-4), Paenibacillus (S1-5), Arthrobacter (S2-1, S2-3, S2-6), Serinicoccus (S2-2, S2-5), Sphingomonas (S2-4), Aureimonas (S2-7), Bacillus (S3-1), Agrococcus (S3-17), Williamisia (S3-18) (Supplementary Table 1). Five additional bacterial isolates (Supplementary Table 2), identified as members of the genera Arthrobacter, Cellulosimicrobium, Sphingomonas, Terribacillus, Bacillus were isolated under microaerophilic conditions.

Characterization of the degradation of chewing gum components

In order to identify possible degrading activities from the chewing gum isolates, a screening was carried out by growing those strains on minimal M9 medium supplemented with different possible components of chewing gum (Fig. 5a). In general, all the isolates grew better in the media supplemented with complete ground chewing gum. The Bacillus altitudinis strain was the only isolate that could grow well in minimal medium alone. Pantoea vagans showed the highest growth when sucrose, mannitol and glycerol were added to the medium as the only carbon source while it grew poorly in the presence of xylitol and sorbitol. Paenibacillus illinoisensis showed a wide spectrum of degradation of chewing gum components except xylitol and sorbitol. In fact, as shown in Fig. 5a, xylitol showed a significant inhibiting effect on the growth of the isolates. Finally, one strain, S2–4, with 98.54% similarity to Sphingomonas insulae, could not grow well on the solid media after isolation. The morphology of some strains was different depending on the type of chewing gum powder that was used as a carbon source for the selective media (Fig. 5b).
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Figure 5

Degradation of various chewing gum ingredients by strains isolated from wasted chewing gums. (a) Heat map representing the ability to grow on different compounds of gum pieces as carbon sources. The phylogenetic tree and taxonomy of the chewed gum isolates based on 16S rRNA gene are shown in the left and right Y-axis, respectively. (b) The strains growth on modified M9 salt media enriched by chewing gum powder I, II, III, compared with the same medium without any carbon source as a control (left).
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Discussion

In this work, we describe the complete characterisation of the wasted chewing gum bacteriome. We report here that wasted chewing gums display a moderate diversity of bacterial population with variations among the samples analysed. Additionally, we show in a specifically designed assay that the oral community-based microbial pool is largely substituted, in a matter of few weeks, by an environmental bacteria-rich microbial community after a process of ecological successions.

The microbial community of the wasted gum samples consists of bacteria from phyla Proteobacteria, Actinobacteria, Deinococcus-Thermus, and Bacteroides. The most abundant families were Sphingomonadaceae, Micrococcaceae, Geodermatophilaceae, and Deinococcaceae and, at the genus level, Sphingomonas, Kocuria, Blastococcus, Deinococcus, and Skermanella; other frequent genera were Nesterenkonia and Hymenobacter. All those taxa have previously been reported as microbial communities associated with natural environments such as the phylloplane27, flowers28 or soil ecosystems29. The taxonomic profile we have detected is similar to abundant phyla on the surface of rocks in Maritime Antarctic glacier forefields30 and also the bacteriome of other outdoor surfaces such as photovoltaic, solar panels. In fact, solar panels are characterised by a desert-like bacteriome rich in Sphingomonas, Deinococcus, or Hymenobacter31. It is obvious that rocks or plant surfaces, solar panels and sun-exposed wasted chewing gums share similar ecological pressures in terms of irradiation, low water availability, thermal variations or oxidative stress.

Streptococcus, Rothia, Haemophilus, Granulicatella, Corynebacterium, Veillonella, Actinomyces, and Gemella were the most frequent genera in the control, chewed sample. These taxa are typical inhabitants of the mouth cavity, which bacteriome is composed of approximately 700 species from 185 genera32, 33. Our results reveal that the pool of oral microbiome bacteria present initially in the chewed samples is substituted after a colonisation process by environmental bacteria. However, some oral genera, specifically Streptococcus, was also detected at relatively high frequencies weeks after being used, as deduced by the colonisation experiment we conducted. Interestingly, Streptococcus was detected with very low relative abundance in most of the older-wasted chewing gum samples we studied, and not detected in samples of Greece and Singapore, suggesting that the stability in time of Streptococcus in the wasted chewing gum may not be more than a few weeks. Furthermore, other oral bacteria Corynebacterium, Haemophilus, Veillonella and Gemella were the most abundant genera in the control sample and these genera remained in the samples through the whole experiment, but their populations clearly decreased over time. The presence of Corynebacterium immediately decreased during the first month of outdoors incubation; however, it was observed with low intensity between 5–12 weeks. Furthermore, the lowest relative abundances from genera Haemophilus, Veillonella and Gemella were detected in sample 12. While, in comparison, the genus Streptococcus was the most frequent in this sample. These results suggest that wasted chewing gums constitute carriers of oral bacteria, some of which could be pathogens, even several weeks after being discarded. The biodiversity of the chewed gum microbial community increased after a few weeks outdoors, and the analysed samples after 6 or 9 weeks were more diverse and richer in genera such as Craurococcus and Sphingomonas, which were found in gum samples until the end of the period analysed. Other genera, such as Cellulomonas and Rubellimicrobium were observed in the gums of the same age as well, but they did not persist in time. Finally, the last samples analysed were rich in Actinobacteria, Blastococcus and other environmental bacteria. A possible explanation for this rearrangement may be that the transient taxa have degraded polymeric substrates remaining in the chewed gum to short carbon chains, and these simple carbon sources could then be hydrolysed by environmental bacteria. Additionally, the changes in abiotic factors such as pH, temperature, oxygen levels or water contents may play a role as selection forces driving community successions.

It is interesting to compare the bacterial profile of the chewing gums exposed outdoors for several weeks in the controlled experiment we performed with that of the “old” wasted chewing gums we sampled and analysed from different locations worldwide. Pseudomonas, Sphingomonas, Rubellimicrobium, and Blastococcus were present as the most abundant genera in almost all samples collected from different countries as well as the last sample of dynamic experiment (sample 12), which indicates that these taxa are fast, common colonisers of the wasted gums. Moreover, other environmental bacteria such as Kocuria, Modestobacter, Deinococcus, and Roseomonas were observed in all samples collected worldwide; however, they only constituted a small percentage of the microbial community in the dynamic samples labelled as “others” (data not shown). This strongly suggests that these taxa correspond to “second wave”, of slower but yet also cosmopolitan, microbial colonisers of the gum substrate.

Wasted gum being compact masses, the access of water and oxygen to the central part of the residues could be at least partially prevented, and it could thus be hypothesized that microbial communities would be different depending on their physical distance to the external environment. As a first approach to find out whether there was a variation in the bacterial composition across the depth of the waste, a single sample was divided into three successive layers that were analysed independently. Surprisingly, no significant differences were observed in the microbial communities of the different chewed gum layers of this sample. This result is striking, since even if the physico-chemical properties of a chewing gum may not change across its depth, UV radiation and water activity would be expected to be strong selective factors shaping a specific bacterial composition. Sphingomonas was detected in all three layers of the analysed chewed gum, as well as Hymenobacter and Deinococcus, all of which have been reported from extreme environments under strong desiccation and radiation conditions34, 35. Due to the limitation of the analysis of a single sample, further research including more samples is needed to address the colonisation of the interior of chewing gums.

Aspartame, mannitol, and glycerol were hydrolysed by several strains that we isolated from wasted chewing gums and identified as belonging to the species Pantoea vagans, Paenibacillus illinoisensis, and Curtobacterium herbarum. Aspartame was also remarkably degraded by Pseudomonas oryzihabitans, Microbacterium arborescens, and Arthrobacter agilis. The biodegradation of several artificial sweeteners including aspartame was recently studied36, 37, but the particular microbial keyplayers involved in the biodegradation process have been poorly studied. Degradation of mannitol and glycerol by Pantoea vagans and Paenibacillus illinoisensis has also previously been reported38, 39. Kuranishi et al.40 proved that Pantoea species can use mannitol as a substrate, by cultivating this genus on a designed semi-selective medium enriched with this compound. Another sweetener, sorbitol, was used as carbon source by one of our strains belonging to the genus Aureimonas. This genus has previously been found to hydrolyse a variety of carbon sources, such as carbohydrates, polyols and organic acids41. Our Curtobacterium strain was able to degrade almost all the tested chewing gum ingredients. Recently, Chase et al.42 reported that Curtobacterium spp. is able to degrade a wide range of carbohydrates, especially structural polysaccharides. It is thus tempting to hypothesize that bioaugmentation of some of the strains we characterized, specially Curtobacterium herbarum, may be used as a bioremediation strategy to contribute to remove chewing gum residues from contaminated pavements.

On the other hand, one of the main compositions of gum base are natural or artificial rubbers43. Previous research reported that these polymers could be degradable by bacteria; nevertheless, the biodegradation of rubbers is a long-term process44 since polymeric chains are cross linked by sulphur binding45. In this study, we have detected, either by culture-dependent or culture-independent techniques some taxa that have been previously described as rubber degraders. Tatangelo et al.46 showed that Rhodococcus can oxidize sulphur to sulphate, and desulphurization can facilitate the rubbers biodegradation. We also detected Bacillus spp. in both culture-dependent and -independent experiments, which has been described as a natural and synthetic rubber degrader44, 47. Interestingly, the population of Bacillus in the control sample of the dynamic experiment was very low; however, the presence of these bacteria gradually increased after several weeks of outdoors incubation. We also identified Sphingomonas spp. as one of the most frequent genera in culture-independent experiments and it was also isolated by culture-dependent methods, which could theoretically have a role in the biodegradation of polycyclic aromatic chains present in the gum base43, 48. Finally, Corynebacterium spp., have been reported as an opportunistic oral bacterium49 as well as a natural rubber degrader that needs direct contact with the surface of rubber particles44. In this research, this genus was detected as a component of the oral microbiome in the control and it remained during 12 weeks in a low-stable frequency that suggests it could play a role in long-term chewing gum biodegradation.

This study is the first report revealing from a holistic approach the bacterial composition of wasted chewing gum. Taken together, our results suggest that bacteria can play a role in the natural biodegradation of the chewing gum and may also be a source of strains with other biodegradation properties. The relative stability of the oral microbiome in a sun-irradiated outdoor space even after several weeks of outdoor and solar exposition raises concerns on the possible role of wasted chewing gums as long-term carriers of pathogenic microorganisms. On the other hand, the fact that the oral microbiome is partially maintained after many days suggest that, besides human DNA analysis, NGS aiming at determining the oral microbiome remaining in a chewing gum could hold potential for legal and forensic applications.
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Material and methods
Chewing gum samples

Wasted chewing gums were collected directly from outdoor pavements in the vicinity of the Scientific Park of the University of Valencia as well from other locations worldwide. In particular, ten wasted gum samples were gathered from five countries (Fig. 1). All samples were removed from the pavement with a sterile scraper, and transported to the lab, where they were kept frozen at − 80 °C until required. In one case, a sample from Valencia (Spain), of approximately 3 mm thick, was sliced in situ in three different layers of roughly 1 mm each (upper, sun-irradiated, dark part; an intermediate part and the bottom fraction of the wasted gum, in contact with the pavement). The resulting three sub-samples were processed independently.

For the assays, two commercial sugar-free chewing gums were used, Orbit and Trident chewing gums. Both chewing gums were ground with a sterilized coffee grinder and they were added to a minimal medium as the sole carbon source. Gum powder I and II were prepared from the same source (Orbit gum pieces). The only difference between those two was that in the gum powder II, the white cortex layer was entirely removed before preparation in order to study whether bacteria only utilize the cortex layer, or they are able to degrade the whole compositions. Gum powder III was prepared from Trident brand.
Media

M9 salt medium (ATCC 2511) was prepared (g l−1): Na2HPO4 12.8, KH2PO4 3.0, NaCl; 0.5, NH4Cl; 1.0, pH adjusted to 7.2. After autoclaving, the following filter-sterilized solutions were added to the medium; 2 ml of 1 M MgSO4 solution, 0.1 ml of 1 M CaCl2 solution, and 20 ml of a Glucose Solution (20% w/v). Glucose as a source of carbon was replaced, when necessary, by possible compositions of the chewing gum. M9 salt medium was modified and enriched with 2% (w/v) gum powder I as a selective medium. In addition to this medium, two rich media, TSA (composition in g l−1: Tryptone 15.0, Soya Peptone 5.0, NaCl 5.0, Agar 15.0) and LB (composition in g l-1: Tryptone 10.0, NaCl 10.0, Yeast Extract 5.0, Agar 15.0), were also used.

In order to study the biodegradation of the chewing gum compositions by isolates, the modified M9 salt medium (without glucose) was enriched with 2% (w/v) carbon sources including; Sucrose, Sorbitol, Mannitol, Xylitol, Aspartame, Gum powder I, II, III, and 2% (v/v) glycerol. Gum powders were used as other possible complementary ingredients of chewing gum.
Culture-dependent approaches

Chewed gums were taken off with a sharp scraper from three locations in an open area from the Scientific Park of the University of Valencia, Spain. Two samples were incubated in liquid M9 salt solution (without glucose) supplemented with 2% (w/v) chewing Gum Powder I, at room temperature orbital shaking (150 rpm) for 24 and 48 h, respectively. The third sample was directly plated on the same solid medium after resuspension. Several serial dilutions were prepared and cultured on the selective media. Plates were incubated at room temperature for a week under aerophilic, anaerobic and microaerophilic conditions. Isolated colonies were selected based on their shape and colour and they were re-streaked on fresh media. Pure isolates were resuspended in 20% glycerol (v/v) and cryo-preserved at − 80 °C.

Polymerase chain reaction was performed with universal primers 8F (5′-AGAGTTTGATCCTGGCTCAG-3′) and 1492R (5′-CGGTTACCTTGTTACGACTT-3′) to amplify 16S rRNA gene. A loopful of bacterial cells was resuspended in 100 µl MilliQ water, pre-incubated at 100 °C for 10 min, and 1 µl of each bacterial suspension was used as DNA template. Thermal cycler program was set as the following procedure; Initial step of incubation at 95 °C for five min followed by PCR amplification (30 cycles of 30 s at 95 °C, 30 s at 54 °C, 30 s at 72 °C), and final step at 72 °C for 10 min. PCR products were monitored by 1% agarose gel electrophoresis to confirm the amplification of the 16S rRNA gene fragment amplicon. Next, dsDNA was purified from the PCR products and resuspended in 10 μl MilliQ water. 16S rRNA Sanger sequencing was performed by tagging with BigDye Terminator v3.1 Cycle Sequencing Kit (Applied Biosystems, Carlsbad, CA, USA), at the Sequencing Service (SCSIE) of the University of Valencia. All sequences were edited and compared with EzBioCloud online database (https://www.ezbiocloud.net/).
Carbon source use

To study the ability of the bacterial strains isolated from wasted chewing gum to degrade different components, chewing gum sweeteners and glycerol were added as carbon sources to the minimal medium (described in media section). Moreover, three different media containing Gum Powder I, II, III separately were prepared to observe its biodegradation. For each isolated strain, a dilution of 0.2 optical density (OD600) was prepared with fresh liquid M9 medium. Then 2 µl of the dilutions were spread on the three different media aforementioned, in three replicates. The plates were incubated at room temperature under aerobic condition for five days. Also, the minimal M9 medium without any carbon source was used as the control medium.
Ecological succession and bacterial colonisation

A study of the microbial colonisation of wasted chewing gum under controlled conditions was carried out. The study protocol was approved by the board of directors of The Institute for Integrative Systems Biology (I2SysBio) and was conducted under the guidelines of Helsinki declaration of 2013. Informed consent of the volunteer was obtained prior the study. A healthy volunteer (36 years old female) chewed two chewing gum pieces every day for 30 min. The first chewed gum was stored in − 80 °C as a control of the oral microbiome. Twelve chewed gums processed this way were placed on the sidewalk of the Scientific Park (University of Valencia, Spain) on an outdoor, sun-oriented location. The experiment was conducted in mid-June, and wasted gums were processed as described for 12 consecutive days. Then, they were picked up in intervals of one week during a total period of twelve weeks. Total DNA was extracted from each sample and 16S rRNA metagenomic analysis was carried out.
16S rRNA sequencing

The following procedures for DNA extraction and 16S rRNA sequencing were performed in all the experiments. All chewing gum residues were frozen after sampling and stored in − 80 °C at least for overnight. Frozen samples were ground to fine powder and added to 1 ml PBS buffer. Mixtures of wasted gum and PBS buffer were frozen at − 20 °C overnight. Then, 10 glass plating beads (3 mm diameter) were added to the tubes and mixed with a vortex for 3 min. Samples were let at room temperature for 2 min, and then 500 µl of the supernatant was transferred to a 2 ml micro-tube. DNA extraction was then carried out as described by Latorre et al. (1987)50, and the extracted DNA was quantified through Qubit dsDNA HS Assay kit (Qubit 2.0 Fluorometer, Q32866).

The conserved regions V3 and V4 (459 bp) of the 16S rRNA gene were then amplified using forward and reverse primers: 5′-TCG TCG GCA GCG TCA GAT GTG TAT AAG AGA CAG CCT ACG GGN GGC WGC AG 3′ and 5′- GTC TCG TGG GCT CGG AGA TGT GTA TAA GAG ACA GGA CTA CHV GGG TAT CTA ATC C -3′, respectively51. Amplification was carried out using the KAPA HiFi HotStart ReadyMix PCR kit (KK2602) and the following PCR cycle: initial denaturation at 95 °C for 3 min; 25 cycles of amplification (30 s at 95 °C, 30 s at 55 °C, 30 s at 72 °C); and 5 min of extension at 72 °C. Amplicons were mixed with Illumina sequencing adaptors and dual-index barcodes (Nextera XT index kit v2, FC-131-2001). Libraries were normalized and merged before the sequencing. Then, the pool containing indexed amplicons was loaded onto the MiSeq reagent cartridge v3 (MS-102-3003), spiked with 10% PhiX control to enhance the quality of the sequencing. Finally, paired-end sequencing (2 × 300 bp) was carried out on the Illumina MiSeq sequencing system. Illumina outcomes were analysed via Qiime2 software52, 53. The Demux plugin was used to assess the quality of reads, and the Qiime2-integrated Dada2 pipeline was employed for trimming and joining the sequences, removing chimeras and detecting sequence variants (> 99.9% of similarity). The classify-Sklearn module (feature-classifier plugin) was applied for assessing the taxonomy of each sequence variant, using the SILVA (v. 132) database as reference.

Data was subsequently analysed by different R packages. Rarefaction curves were constructed with the iNEXT and ggiNEXT functions (iNEXT)54. Alpha diversity plots were generated by employing the plot_richness function (Phyloseq)55 based on the richness, Shannon and Simpson diversity indexes. PCoA were created with the plot_ordination function (phyloseq) using Bray–Curtis dissimilarities as distance method. Finally, heatmap was constructed by the heatmap.2 function (gplots).
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Supplementary information

Supplementary file1(146K, pdf)

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Acknowledgements

Financial support from the Spanish Government on SETH Project (Reference: RTI2018-095584-B-C41-42-43-44 co-financed by FEDER funds and Ministerio de Ciencia, Innovación y Universidades) and from the European CSA on biological standardization BIOROBOOST (EU Grant number 820699) are acknowledged. LS is funded by European project BIOROBOOST. ÀVV is funded with a FPU (Formación de Profesorado Universitario) grant from the Spanish Government (Ministerio de Ciencia, Innovación y Universidades), with reference FPU18/02578. We are in debt with Adriel Latorre for his assistance with the bioinformatics analysis.
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Author contributions

M.P. designed the work, L.S. and À.V.V. carried out the experiments, A.G. and L.S. carried the analysis, and all authors wrote and approved this manuscript.
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Data availability

Raw reads are available at NCBI’s Sequence Read Archive (SRA) (Bioproject Accession PRJNA641111).
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Competing interests

The authors declare no competing interests.

[AsbestosSniffer]

Uh? Starsector?

[ariever]

I cant play ff14 till thursday, why even live

[Qbmax32]

lol

[MrStonedOne]

firstly, this thread sucks.

secondly, these discussions always hide some inner political belief that people are subtly deferring to.

as usual i will use my personal life experience to tell a story about this:

i'm a woman and when i was younger i spent a lot of time online and wanted to be liked by my peers and accepted into "the culture" of 2008 era 4chan shitposting. i would tell intolerant jokes, say bigoted things and mock "normies" because they were the outside and i needed social cred to elevate myself on the inside.

years later in my late teen years i started hanging out with other women who were politically active and they taught me about feminism and i had a phase where i was one of those "men are the problem" types and i was just as much of an insufferable jackass but now i hung out on the other side of the divide.

then a few years later, in my early-mid 20s, i had calmed down because i had made friends with men, women, normies and internet dwellers all alike. i started to find it difficult to tell those same jokes, to hold those same beliefs against men or women because every time i did i would think of my friends, their life experiences, the problems they faced and i knew it would hurt them to hear me say those things and it became impossible to be so reductive to the true breadth and depth of their struggles and experiences. i care about them, no matter who they are, and i want to help them self actualize and be free from the things causing them pain.

what i'm ultimately trying to say is that it does no side any good to paint the other with a broad brush. i still believe in intersectionality: the idea that all people are a big ball of different types of privilege and discrimination based on a wide array of attributes. men wield the majority of power in society, but that power does not benefit all men. access to mental healthcare is worse for men, suicide affects them more, it is harder for them to show emotional vulnerability, they are the ones who are made to take dangerous jobs, they are the ones that die in wars, they are the ones who often lose access to their children in divorce, the list goes on and on. obviously it goes without saying that there is a whole host of issues that affect women too. it doesn't mean any of that stops existing like it's some sort of competition over who gets fucked over more. we all do. the thing that will change you and better you in this regard is to make friends with everyone, even those you might disagree with. they will see you as a real person, not some political opposition that's dangerous to them, and you will see the same in them too, and both of you will grow from sharing your experiences and your struggles.

wtf dendy is now my new favorite admin? (sorry arm)

[toemas]

garbage

You're incoherently regurgitating a bunch of retarded liberal platitudes that have no relevance. Men commit more violence than women because they're biologically stronger than women, and they're mentally inclined to engage in territorial/aggressive behavior. This is the case for almost every species of animal that exists on the earth.

[Super Aggro Crag]

firstly, this thread sucks.

secondly, these discussions always hide some inner political belief that people are subtly deferring to.

as usual i will use my personal life experience to tell a story about this:

i'm a woman and when i was younger i spent a lot of time online and wanted to be liked by my peers and accepted into "the culture" of 2008 era 4chan shitposting. i would tell intolerant jokes, say bigoted things and mock "normies" because they were the outside and i needed social cred to elevate myself on the inside.

years later in my late teen years i started hanging out with other women who were politically active and they taught me about feminism and i had a phase where i was one of those "men are the problem" types and i was just as much of an insufferable jackass but now i hung out on the other side of the divide.

then a few years later, in my early-mid 20s, i had calmed down because i had made friends with men, women, normies and internet dwellers all alike. i started to find it difficult to tell those same jokes, to hold those same beliefs against men or women because every time i did i would think of my friends, their life experiences, the problems they faced and i knew it would hurt them to hear me say those things and it became impossible to be so reductive to the true breadth and depth of their struggles and experiences. i care about them, no matter who they are, and i want to help them self actualize and be free from the things causing them pain.

what i'm ultimately trying to say is that it does no side any good to paint the other with a broad brush. i still believe in intersectionality: the idea that all people are a big ball of different types of privilege and discrimination based on a wide array of attributes. men wield the majority of power in society, but that power does not benefit all men. access to mental healthcare is worse for men, suicide affects them more, it is harder for them to show emotional vulnerability, they are the ones who are made to take dangerous jobs, they are the ones that die in wars, they are the ones who often lose access to their children in divorce, the list goes on and on. obviously it goes without saying that there is a whole host of issues that affect women too. it doesn't mean any of that stops existing like it's some sort of competition over who gets fucked over more. we all do. the thing that will change you and better you in this regard is to make friends with everyone, even those you might disagree with. they will see you as a real person, not some political opposition that's dangerous to them, and you will see the same in them too, and both of you will grow from sharing your experiences and your struggles.

wtf dendy is now my new favorite admin? (sorry arm)

before you declare her your favorite admin please be aware she is ENGLISH

[AsbestosSniffer]

snip

wtf dendy is now my new favorite admin? (sorry arm)

before you declare her your favorite admin please be aware she is ENGLISH

Our power grows.

[Lacran]

garbage

Retarded liberal

But size and behaviour are not causal or uniform, neither are the actual differences in statistics between genders.

Populations can have larger men with more testosterone yet less violent crime meaning that these just correlate and aren't why this actually happens.

It's easy to say men and women have different statistics in crime due to biology. But it's way too simplistic and ignores why crime actually occurs and increases/decreases within a population.

I don't wanna drag this convo out here though, ping me on the tg discord in politics if you wanna discuss this more.

[EmpressMaia]

what the fuck happeed to this thread. go touch grass, all of you

[AsbestosSniffer]

what the fuck happeed to this thread. go touch grass, all of you

what if they live in the desert

[Fren256]

the thing that will change you and better you in this regard is to make friends with everyone, even those you might disagree with. they will see you as a real person, not some political opposition that's dangerous to them, and you will see the same in them too, and both of you will grow from sharing your experiences and your struggles.

See guys this is why we should befriend neo-nazis

[toemas]

But size and behaviour are not causal or uniform, neither are the actual differences in statistics between genders.

Size and behavior are absolutely "causal" and the rest of this is just irrelevant

Populations can have larger men with more testosterone yet less violent crime meaning that these just correlate and aren't why this actually happens.

This is like saying that poverty doesn't cause crime because "Some poor populations commit less crime than some rich populations" Completely incoherent logic

It's easy to say men and women have different statistics in crime due to biology. But it's way too simplistic and ignores why crime actually occurs and increases/decreases within a population.

the statement "Men commit more crime than women because they're biologically inclined towards violence and aggression" is indisputably true, the existence of various socio-economic factors that result in crime don't change the reality of this

I don't wanna drag this convo out here though, ping me on the tg discord in politics if you wanna discuss this more.

I'm not going to do that, I'm not in the discord.