My heartfelt thanks go out to all of you who have supported the Children's Place Association in 2015 through your gifts and volunteer efforts. Your kindess and generosity made an enormous difference in the lives of so many children and families who are overwhelmed by illness and poverty.
Don't forget, you still have time to make your year end 100% tax-deductible donation to the Children's Place. Even a small donation can make a difference.
Thank you again for your unwavering support, you are truly a CHAMPION for Chicago's most vulnerable children.
Best wishes for a happy, healthy New Year.


text:
Patiala MP Maharani Preneet Kaur Faints During Cleanup Campaign
Jul. 13 Environment, Glamour, Politics Comments
[Patiala,Pb]Patiala MP Maharani Preneet Kaur Faints During Cleanup Campaign
Patiala MP Preneet Kaur fainted at a plastic cleanup campaign in her constituency
The Congress MP and wife of Punjab Chief Minister Amarinder Singh collapsed as she arrived to flag off the Swachhta Shramdan drive to clean up plastic.
The 75-year-old, later, on her Facebook page said she successfully launched the drive under the Punjab Pollution Control Board.
PM Modi's One Dayer Varanasi Visit Results Three Programme
Jul. 06 Environment, Politics, Social Cause Comments
[Varanasi,UP]PM Modi's One Dayer Varanasi Visit Results Three Programme Of Govt+Party+Environment
Prime Minister Narendra Modi unveiled 18 feet high statue of former PM Lal Bahadur Shastri on Saturday during his day-long visit to his Lok Sabha constituency .
Modi was accompanied by Uttar Pradesh Chief Minister Yogi Adityanath, Bharatiya Janata Party (BJP) working president JP Nadda and the state unit chief of the saffron party, Mahendra Nath Pandey.
Anil Shastri and Sunil Shastri, sons of late Lal Bahadur Shastri, were present at the venue as also Uttar Pradesh minister Siddharth Nath Singh, a kin of the former prime minister.
Earlier, Modi arrived at the Babatpur airport here on his second visit to his constituency after winning the parliamentary polls held in April-May.
The prime minister launched a tree-plantation campaign — "Anand Kanan" — in the temple city, he planted a Peepal [Sacred Fig Tree ] PM Modi Also kickstarted the membership drive of the ruling party, coinciding with the 118th birth anniversary of Bharatiya Jana Sangh founder Syama Prasad Mookerjee.
HC Agrees to Hear PIL Against Population Explosion in India
May. 28 Economy, Environment, Social Cause Comments
[New Delhi] HC Agrees to Hear PIL Against Population Explosion in India
The Delhi High Court on Tuesday agreed to hear a PIL seeking directions to the Centre to implement measures to control the country's rising population
The plea by BJP leader Ashwini Kumar Upadhyay, who is also a lawyer, has sought implementation of the recommendations for population control made by the National Commission to Review the Working of the Constitution (NCRWC) headed by Justice Venkatchaliah.
"The NCRWC, after making immense efforts for two years and elaborate discussion, had suggested addition of Article 47A in the Constitution and formulation of Population Control Law.
"Till now the Constitution has been amended 125 times,
The plea claims that the population of India has "marched ahead" of China, as about 20 per cent the population does not have Aadhaar and therefore, is not accounted for, and there are also crores of Rohingyas and Bangladeshis living illegally in the country.
ISRO Decorated with Another Feather By Launching RISAT-2B
May. 22 Environment, Glamour, World Comments
[ISriharikota] SRO Decorated with Another Feather By Launching RISAT-2B
India's Polar Satellite Launch Vehicle (PSLV-C46) today successfully launched the RISAT-2B satellite from Satish Dhawan Space Centre (SDSC) SHAR, Sriharikota in Andhra Pradesh. This was the 72nd launch vehicle mission from SDSC SHAR, Sriharikota and 36th launch from the First Launch pad.
RISAT-2B is a radar imaging earth observation satellite weighing about 615 kg. The satellite is intended to provide services in the field of Agriculture, Forestry and Disaster Management.
A total number of 5,000 visitors witnessed the launch live from the Viewer's Gallery, which is opened to the public.
ISRO is now gearing up for the launch of Chandrayaan-2 onboard GSLV MkIII during the window of July 09, to July 16, 2019, with an expected Moon landing on September 06, 2019.
Earth Quake Jolts Its Own Favourite Andaman & Nicobar With 5.8-Magnitude
May. 22 Environment Comments
[New Delhi]Quake Hits Andaman & Nicobar With 5.8-Magnitude
A medium intensity earthquake of magnitude 5.8 hit the Andaman and Nicobar islands on Wednesday morning.
As per National Centre for Seismology,This quake, had its epicentre in the Andaman islands,Which occurred at 6:09 am,
The Andaman and Nicobar archipelago is prone to earthquakes.
200,000 Zimbabweans Affected by Floods+Cyclone
Mar. 21 Environment, Social Cause, World Comments
[Delhi,Geneva]200,000 Zimbabweans Affected by Floods+Cyclone
The number of people in Zimbabwe affected by a devastating cyclone and flooding has jumped to 200,000
The initial estimate of those hit in Zimbabwe was 15,000 but World Food Programme (WFP) spokesman Herve Verhoosel told reporters in Geneva that the numbers had surged following an overnight assessment.
Five days after tropical cyclone Idai cut a swathe through Mozambique, Zimbabwe and Malawi, the full extent of the devastation is not yet clear.
In hardest-hit Mozambique, an estimated 400,000 people have been displaced, according to the UN.
राजस्थान में मंगलवार की सुबह चार राष्ट्रीय पक्षी मोरों[पीकॉक] के शव बरामद
Mar. 19 Crime, Environment Comments
[जयपुर]राजस्थान में मंगलवार की सुबह चार राष्ट्रीय पक्षी मोरों[पीकॉक] के शव बरामद
|मोर राष्ट्रीय पक्षी है और वाइल्ड लाइफ प्रोटेक्शन एक्ट के अंतर्गत इसे सरंक्षण मिला हुआ है |
राजस्थान के भीलवाड़ा में खाखरमाला गावं में आज सुबह सरकारी भूमि पर चार मोर मृत मिले
बीते वर्ष बूंदी के नैनवा में २५० मोर मार डाले गए थे|प्राप्त जानकारी के अनुसार पक्षी तस्कर इस राष्ट्रीय पक्षी का शिकार करके इसके पंख और मीट की तस्करी करते हैं
India to Provide Disaster Relief Support to Mozambique
[New Delhi] India to Provide Disaster Relief Support to Cyclone Hit Mozambique on Humanitarian Grounds
Government of India has decided to divert three Indian Naval Ships
INS Sujatha,
INS Shardul &
INS Sarathi
to the port city of Beira to provide immediate Humanitarian Assistance and Disaster Relief (HADR) to the affected people.
Indian Naval Ships will provide relief material in the form of
clothes and
medicine to the affected people.
In addition, the ships have 3 medical practitioners and 5 nurses to provide immediate medical help.
In this hour of tragedy, Government of India stands ready to extend support to the affected people. India has been extending humanitarian assistance to Mozambique and had provided 10 million dollars for food grains in 2017, after it suffered food shortage as a result of natural calamities.
Kamal Nath Blesses Computer Baba With Chairmanship Of River Trust in MP
Mar. 12 Environment, Politics Comments
[Bhopal]Kamal Nath Blesses Computer Baba With Chairmanship Of River Trust in MP
The Congress government in Madhya Pradesh has appointed self-styled godman NamdevTyagi, also known as Computer Baba, as chairman of a river trust in the state
The state spiritual department passed the order of appointing Tyagi as head of the 'Ma Narmada, Ma Kshipra and Ma Mandakini Rivers Trust' a few days back, just before the model code of conduct for Lok Sabha polls came into effect
Notably, Tyagi was accorded minister of state (MoS) status by the previous BJP-led government last year, but he later resigned, accusing the then chief minister Shivraj Singh Chouhan of "not fulfilling" the promise of stopping illegal sand mining in the river Narmada.
Tyagi, a friend-turned-critic of Chouhan, campaigned against the BJP ahead of the last year's state Assembly polls in which the Congress dethroned the saffron party and came to power after over 15 years
Capt's Cabinet Gives Green Signal To Brick klins in Punjab
Mar. 06 Economy, Environment Comments
[Chd,Pb] Capt's Cabinet Gives Green Signal To Brick klins in Punjab
This approval is for rectangular shape and zigzag brick setting by Punjab State Council for Science & Technology. Brick kilns have been given till September 30, 2019, to make the conversion.
The brick kilns in the process of conversion to induce draft technology with zigzag setting/yet to start the process intending to operate the brick kiln are required to pay environmental compensation on the basis of `polluter pays principle' to the Punjab Pollution Control Board (PPCB) with effect from February 1, 2019, till the adoption of induced draft technology. The compensation has been set Rs. 25,000 per month for zig zag setting with kiln of capacity equal to more than 30,000 bricks per day and Rs. 20,000 per month for kiln of capacity less than 30,000 bricks per day.
The PPCB would ensure that the brick kilns are operated in compliance with the code of practice prescribed by the CPCB, as well as the state board. The Department of Food and Civil Supplies, being the licensing authority, would create awareness and mobilise the brick kiln for conversion to induced draft technology with zigzag setting.
482 brick kilns had already converted to new technology and 559 were in pipeline for shifting to induced draft technology.
Notably, around 2800 brick kilns are operating in Punjab, producing around 15-20 billion bricks per annum. This constitutes about 8% of total production of the country and the brick kiln industry employs about 0.5-0.6 million workers.

redpajama_set_name: RedPajamaCommonCrawl


text:
Security stepped up as India-Pakistan clash nears
Indian all-rounder Yuvraj Singh plays football at a practice session ahead of the World Cup semifinal against Pakistan in Mohali on Sunday. (AP)
Published Monday, March 28, 2011
Security was ramped up at the Punjab Cricket Association stadium on Sunday as India staged their first training session ahead of the high-profile World Cup semifinal against Pakistan.
The match, the first between the arch-rivals on Indian soil since the 2008 Mumbai terror attacks, gained an added security headache on Sunday when Pakistan's Prime Minister Yousuf Raza Gilani confirmed he will attend Wednesday's clash.
Police were out in greater numbers in and around the ground on Sunday and there were even "media security officers" escorting reporters into the ground.
Pakistan, the 1992 champions, were put through their paces once again by coach Waqar Younis in an early morning training stint that started with a lively football match.
Coming back to the PCA Stadium was proving particularly pleasant for Pakistan manager Intikhab Alam, who earlier in his career coached a Punjab team featuring both Yuvraj and India off-spinner Harbhajan Singh.
"I am nostalgic, I remember this place very well," former Test leg-spinner Alam told the Sunday Pioneer.
"The two years I spent here earned me friends for life. It feels great to come back."
The India squad arrived for their scheduled training session later in the day, with off-spinner Harbhajan Singh the only notable absentee.
Like Pakistan, the co-hosts too began with a potentially injury-inducing game of football.
However, it was notable how Sachin Tendulkar, who comes into this game on the back of 99 hundreds, stood away from the fray and opted for the more traditional warm-up method of knocking in a few bats.
Meanwhile flag-waving 'super-fan' Sudhir Gautam from Bihar, eastern India, was the lone supporter joining the massed ranks of reporters, photographers and television crews watching training in the stands.
Around 3,000 police will patrol Wednesday's match with some 2,000 expected to be deployed in and around the 30,000-capacity PCA Stadium on matchday.
An estimated 1,000 police have already descended on the luxury Hotel Taj in nearby Chandigarh where both teams are staying, a force which includes Indian army commandos.
The game, which takes place in the border state of Punjab, has already sparked a flurry of political activity with confirmation that Gilani has accepted an Indian government invitation to watch the match.
Pakistan batsman Misbah-ul-Haq welcomed the move, saying: "It's a good thing, no pressure. He's helping to support us.
"And I think it's a good gesture from the Indian Prime Minister that he invites him to come here and watch the game."
Elsewhere the frantic scramble for hotel rooms and tickets showed no signs of slackening, even though the PCA had insisted as early as Tuesday that the 14,000 available tickets had been sold, with the rest in the hands of the ICC.
However, there have been numerous reports of a thriving black market in tickets with prices rocketing so that a 5,000-rupee ($112) ticket was being sold for as much as 25,000 rupees ($560).
"For a match as big as this one, we cannot do anything about black market tickets," said PCA joint secretary GS Walia.
"As far as we are concerned, we only gave the tickets to those who stood in the queues and bought them. We cannot keep track if they sell these in (the) black (market)."


The rulers are on an enterprise of conquest. They take whatever they can from whoever has anything using any means at their disposal. Violence and destruction accompany their efforts as surely as bullets a gun. This is as obvious as the flow of money (to the rulers) and the spread of conflicts and despair to everyone else (the people).
Opposing the rulers is a struggle for survival. The rulers must be opposed, not appealed to or influenced. The people must be helped, supported, encouraged, educated and united.
The rulers dominate the public sphere. They rule it. The people must act among the people to form the force that can weaken, weaken, weaken and defeat the rulers. Weaken the enemy and strengthen your own forces. That's as basic as it gets. The rulers are also subject to these basics, but we don't have to play by their rules.
"Play by life's rules or don't play at all.


Bill Frisell ventures to Nashville to explore Americana with a supporting cast of Music City greats, including dobro virtuoso Jerry Douglas and bassist Viktor Krauss, and guest vocals from Robin Holcomb. "Comprising acoustic instrumental folk tunes with unpredictable stylistic accents," L.A. Weekly declared, "Nashville boasts a dreamy, seductive grandeur." This edition was mastered for vinyl by Greg Calbi at Sterling Sound and pressed in a two-LP set on 180g vinyl at Record Industry in the Netherlands. The set comes in a gatefold tip-on jacket made at Stoughton Printing with outtake photos from the sessions.


text:
extern "C" {
#include "libavutil/opt.h"
#include "libavutil/imgutils.h"
#include "libavformat/avformat.h"
#include "libswscale/swscale.h" // for BGRA to YUV420p conversion
}

namespace gg
{

class VideoTargetFFmpeg : public IVideoTarget
{
protected:
    //!
    //! \brief \c finalise() proceeds only if
    //! this is \c true, due to Issue #130
    //! \sa finalise
    //!
    bool _is_finaliseable;

    //!
    //! \brief
    //!
    std::string _filepath;

    //!
    //! \brief
    //!
    AVFormatContext *_format_context;

    //!
    //! \brief
    //!
    AVStream * _stream;

    //!
    //! \brief
    //!
    std::string _codec_name;

    //!
    //! \brief For fetching FFmpeg's HEVC encoder using
    //! x265
    //!
    static const std::string _CODEC_NAME_HEVC_X265;

    //!
    //! \brief For fetching FFmpeg's HEVC encoder using
    //! kvazaar
    //!
    static const std::string _CODEC_NAME_HEVC_KVAZAAR;

    //!
    //! \brief For fetching FFmpeg's HEVC encoder using
    //! NVIDIA VIDEO CODEC SDK
    //!
    static const std::string _CODEC_NAME_HEVC_NVENC;

    //!
    //! \brief For fetching FFmpeg's VP9 encoder using
    //! libvpx
    //!
    static const std::string _CODEC_NAME_VP9_LIBVPX;

    //!
    //! \brief
    //!
    AVCodec * _codec;

    //!
    //! \brief
    //!
    AVFrame * _frame;

    //!
    //! \brief Pixel depth (bits-per-pixel)
    //!
    int _pixel_depth;

    //!
    //! \brief Flag indicating first frame being
    //! appended, i.e. to infer the frame params
    //! like width and height automatically
    //!
    bool _first_frame;

    //!
    //! \brief
    //!
    int64_t _frame_index;

    //!
    //! \brief
    //!
    float _framerate;

    //!
    //! \brief Packet for use when encoding and
    //! subsequently writing frames
    //!
    AVPacket _packet;

    //!
    //! \brief This is for converting pixel format
    //! (e.g. from BGRA to YUV420p)
    //!
    SwsContext * _sws_context;

    //!
    //! \brief
    //! \sa _sws_context
    //!
    const uint8_t * _src_data_ptr[1];

    //!
    //! \brief
    //! \sa _sws_context
    //!
    int _stride[1];

public:
    //!
    //! \brief Constructor defining what \c codec to
    //! use for encoding
    //! \param codec `HEVC` or `VP9`
    //! \param filename
    //! \param frame_rate
    //! \throw VideoTargetError with a detailed error message
    //! if \c fourcc not supported or if the target cannot be
    //! initialised for some reason
    //!
    VideoTargetFFmpeg(const std::string codec,
                      const std::string filename,
                      const float frame_rate);

    //!
    //! \brief
    //!
    virtual ~VideoTargetFFmpeg();

public:
    void append(const VideoFrame & frame);

protected:
    void init(const std::string filepath, const float framerate);

    void finalise();

protected:
    //!
    //! \brief Prepare frame data for encoding
    //! \param data
    //! \param data_length
    //! \param width
    //! \param height
    //! \param colour_space
    //! \param frame
    //! \sa encode_and_write
    //! \throw VideoTargetError with a detailed
    //! message on failure in any step
    //!
    void ffmpeg_frame(const unsigned char * data,
                      const size_t data_length,
                      const int width, const int height,
                      const AVPixelFormat colour_space,
                      AVFrame * frame);

    //!
    //! \brief Convenience function for code reuse when
    //! writing delayed frames
    //! \param frame leave \c NULL if writing delayed
    //! frames
    //! \param got_output used as \c got_packet_ptr
    //! parameter of \c avcodec_encode_video2() function
    //! of FFmpeg
    //! \throw VideoTargetError propagates any exception
    //! to caller
    //! \sa finalise
    //!
    void encode_and_write(AVFrame * frame, int & got_output);

protected:
    DISALLOW_COPY_AND_ASSIGNMENT(VideoTargetFFmpeg);
};

}

redpajama_set_name: RedPajamaGithub


text:
Bristol Dig Berkeley
This blog details the University of Bristol's annual archaeological excavations in the grounds of Berkeley Castle, Gloucestershire.
Third Years' Final Dig: Memories of Dig Berkeley
For third years, the dig is no longer a compulsory part of their degree. Those who have returned to the dig this year do so because of their love for the project, and the great memories associated with it. As we begin the third week of excavation, we asked third year students to reflect on their time at the Dig Berkeley project, and share some of these memories. Having been involved in the project over the last three years, they had a lot to share, from favourite memories to most challenging tasks.
Left to right: Dan, Hattie and Max
Public Engagement Officer Hattie says her best memory of the project was getting to bond with people over hard work; coincidentally she said that she found this most challenging but at the same time it strengthened friendships. Hattie says Berkeley is a great site to work at because of its many layers of occupation, meaning there is always something new and exciting-or, as Hattie put it, "old and exciting"-to be found. Managing the project's Public Engagement team has developed Hattie's love for making archaeology accessible; she believes "heritage belongs to everyone". For Hattie, Berkeley will always be the place which cemented her passion for archaeology.
Volunteer digger Dan says he has enjoyed digging at Berkeley because he likes working outside, particularly in the countryside. He says his favourite memory was of getting a bit too much sun in his first year of excavation, and not being able to stop laughing all afternoon, while his biggest challenge has been learning to remember all the archaeological terminology. Dan says he feels lucky to have worked at Berkeley because there is so much archaeology; no matter where you dig, you know you will always find something! He also likes being able to come to such a beautiful part of the countryside, which is home to a historic which contains many period buildings. Dan says being part of Dig Berkeley has "been a real journey". To Dan, Berkeley means contentment: "I feel fuller here than in Bristol".
Assistant Dig Supervisor Max reflected on the three years he spent as part of the Dig Berkeley project, describing it as a fulfilling experience which has left him feeling prepared for the world of professional archaeology. He says his best memory was when he took part in the GPR survey that was carried out inside the Church of St Mary on the Castle grounds. As a result, he managed to develop a deeper understanding of surveying skills. Max enjoys being able to work at Berkeley because it covers a wide variety of eras, such as Roman, Norman and Tudor. He also enjoys when an occasional modern day find is discovered on site. Max thinks that the most important thing that he has taken away from his time at Dig Berkeley is preparation for the work place. He praises the dig as a training excavation which provides its students with "the foundation for future careers". For Max, Berkeley means getting to do good work with good people.
A Guide to Archaeological Illustration
Spotlight on Post-Excavation: Where Do All The Fin...
Slag: Not Just Waste
Who Is Your Archaeological Hero?
The Next Generation of Archaeologists
What Time Period Do You Belong In?
A Typical Norman Town
Springs, Pubs and a Mystery Feature...
Berkeley Castle Exhibition: Our latest ambitious p...
GNSS and its Uses at Berkeley
Animal Bones Galore!
Which Famous Archaeologist Are You? Take Our Quiz!
Back at Berkeley: 14th Time Lucky


Комментируем особенности моделей и карт для OMSI.
Норм автобус.Порадовала регулируемая приборка и звуки.
You can fix it easily! Go to folder "Texture" and find 3 files with name "knöpfe_.jpg" e.g. Change this to "knцpfe_.jpg" and will work.
On my system it works. Changе ö to cyrilic "ц" and ü to "ь".
Other option is to open model.cfg and try to find "knopfe..." texture rows and change the deutsch symbols with normal.
That german people didn't want to make buses for normal people I suppose. That's the n-bus with german letters in files.


text:
2020 Mini Cooper SE revealed
Mini shows off the upcoming electric Cooper SE
Mini has revealed the Cooper SE, its first ever series-production electric vehicle.
Power in the electric Mini comes from a 135kW/270Nm electric motor on the front axle, allowing the three-door hatch to accelerate from 0-60km/h in 3.9 seconds and 0-100 in 7.3 seconds. Top speed, meanwhile, is limited to 150km/h.
The motor is fed by a 32.6kWh lithium-ion battery pack which the company says is 'model specific', allowing for a claimed driving range of 235 to 270 kilometres.
Both AC and DC charging methods are supported by the Cooper SE, meaning it is capable of 50kW fast charging which replenishes 80 per cent of the battery in 35 minutes.
Compared to the combustion-powered 3 Door Hatch, the Cooper SE rides 18mm higher to accommodate the battery pack mounted under the vehicle's floor, while the EV also sports unique design elements inspired by the Mini Electric concept.
Despite the added height, the company claims the Cooper SE has a centre of gravity "at least 30 millimetres lower" than the equivalent Cooper S, which should ensure that 'go kart feel' the brand is known for.
Luggage capacity hasn't been impacted, either, with the Cooper SE claiming the same 211L capacity with the second row in place and 731L with the rear backrests folded down.
Mini adds the Cooper SE is around 145kg heavier than the equivalent Cooper S 3 Door, at 1365kg unladen.
Standard equipment includes 16-inch light alloy wheels, a 6.5-inch Connected Navigation system, LED headlights with Union Jack LED tail-lights, dual-zone climate control, fabric seat trim, 'Vigorous Grey' mirror caps, and compatibility with the Mini Connected Remote App which allows for Remote Services like auxiliary heating, vehicle data and more to be accessed via a smartphone. There's a unique digital driver's display and other model-specific elements in the interior, too.
In the UK, the Mini Cooper SE is available to order now priced from £24,400 ($43,883), with deliveries to commence in March 2020.
Mini's local division says it is unable to confirm local availability or timing for the Mini Cooper SE at this stage.
2020 Mini Cooper SE
James Wong
DJM61 — 11 Jul 2019 11:10
a claimed driving range of 235 to 270 kilometres ... it won't be coming here?
(2) Comments on: 2020 Mini Cooper SE revealed
2020-mini-cooper-se-revealed-121891
['16938419','13910441','16938437','17146117']


text:
Q: April Fool's Day changes should be based on the user's time zone No, this isn't a duplicate of all the normal "make badges and the rep cap day local" requests. It's very specific to April 1st.
Users should be required to specify their time zone next time they log in to Stack Overflow:

*

*Don't try to guess it from JavaScript. That will be ambiguous.

*Don't present a list; lazy users will just pick a random entry

*Insist on both the Windows system ID for the time zone and the Olsen database name. That way there's no room for ambiguity.

*If a user can't get a valid combination after 3 attempts, suspend them. They're clearly not trying hard enough.

*Allow a user to change their time zone only when the date of their current time zone is the same as the date within the new time zone.

*Periodically confirm their time zone by giving them  pop quiz on when their DST will next change (if at all).

Then, and only then, can we make sure that any April Fool's Day specific features show up at the right time.

A: I disagree.
I'm Israeli, we set DST at random intervals according to local politics, and I will sure to fail that test.
The jokes belong on Purim, Adar 14 according to the Hebrew calendar, and Adar 15 on ancient cities surrounded by walls. On leap years, the Jewish calendar has another month - Adar A and Adar B, and Purim is held on Adar B. Of course, if that falls on a Sabbath, the holiday can be postponed by a day.
Now, implement that.

A: To make it complete we should also make sure we collect the cultural preferences for the users, to properly present special features on the correct local fools' day, According to Wikipedia there are a number of other days to take into account:

*

*1st or 2nd of April in Iran, depending on exactly when the Persian new year occurs.

*December 28th in many Spanish-speaking countries (should be the default for a Spanish version of SO).

*1st of May in Denmark and southernmost Sweden.

*The Israeli rules according to @Kobi's post.

A: Why limit it to the current day? Users viewing historical questions might get confused when their time zone doesn't match the time zones in effect when the question was asked. Using the historical tzdata information as a reference, require the user to correctly distinguish "summer time", "war time", "United States standard eastern time", and the like; and to correctly figure out what time it is on any given day in arbitrary cities of Indiana.

A: Seconded. Otherwise, the site could be in April Fool's mode for hours after it has already ended in the user's time zone. TIt would reflect poorly on Stack Overflow's professionalism.
The pop quiz would have to be really quick (like, ten seconds) to prevent cheating. Also, it needs some blinking lights and sound.
Also, thanks @jleedev, only one spelling of each time zone name can be allowed. Every deviation must result in a ban.

A: I decry all of this cultural relativism.  If internet history teaches us anything, it is that April Fool's Day begins whenever it is April 1st anywhere and that the worst faux pas is to release an April Fool's Day joke early (releasing it late is just lame).  Due to the spotty international connectivity of the early internet, there is a mistaken belief that it must be April Fool's Day on the east coast of the US.  Due to this belief, it is sometimes considered safest, to avoid the cardinal sin of releasing the joke early, to wait until 12:00AM ET before unleashing the hounds.  Informed netizens know, however, that as soon as it hit's April 1st on the International Date Line the jokes are acceptable.
On a related note, I understand that they've recently replaced kremvax with a quad-core PC running Ubuntu Server-Edition and are bringing up a StackOverflow clone on it.

redpajama_set_name: RedPajamaStackExchange


Are you looking for safe and effective thermogenic supplements for weight loss?
Enriched with green tea catechins, yohimbine, and caffeine, this thermogenic supplement contains 7 powerful ingredients which are known to boost the energy levels and accelerate fat loss. The supplement is vegan-friendly and gluten-free. It works by enhancing the metabolism of the body and defining the muscles to give you a slimmer appearance. The supplement also works on improving the cognitive functions. The green tea extracts reduce excess fat in the body and control cravings while the other ingredients work on the energy levels and protect the muscles. The supplement also contains gelatine so make sure that your body is engaged in some mild form of exercise daily to burn the extra energy. Mild side-effects like an uneasy stomach and lack of sleep can be seen in some users but they go with time as you get accustomed to the formula. Each pack contains 60 capsules which are supposed to last for 30 days. The price of the supplement is moderate as compared to other supplements.
This thermogenic supplement is good for people who want to lose weight naturally over a period of time. It gives slow but steady results with its green tea and caffeine- enriched formula.
Garcinia Cambogia is a well-known ingredient for burning excess fat in the body and this supplement from Zenutra is made using Garcinia Cambogia, green coffee extracts, and raspberry ketones. This combination burns the fat by increasing the heat in the body and boosting the energy to enable the muscles to work harder. The formula also contains African mango which is known to preserve the muscles and reduce cravings. It does have a slightly strong aftertaste which takes some time to adjust to but the results are very good. Regular use will give you a slimmer and muscular body without any harmful side-effects like the case with other thermogenic supplements. All the ingredients used in this supplement are natural and herbal. The packaging is compact and travel-friendly so you do not have to miss taking your supplement even when you are out on a tour or a vacation. The price of the supplement is a bit higher than other brands.
Some people find Garcinia Cambogia a better ingredient for weight loss and this supplement uses that ingredient in a safe and effective manner along with other fat-burning constituents. The natural formula and the results make this supplement worth the price.
Designed to suppress cravings and enhance energy and focus, this supplement uses the fat-burning properties of black pepper and green tea to make you lose weight faster. Each bottle contains 60 veggie capsules which are supposed to be taken for a month with 2 capsules for each day. The supplement does not have any side-effects like increased heart rate, rashes, or jitters. The caffeine and cayenne pepper extracts work in boosting the blood circulation in the body and increasing the heat so that the muscles burn more fat in lesser time. They also enhance the sweating in the body so that it can cool down faster after a workout. It also improves the focus and concentration of the mind which helps the body with the healing process. Make sure that you balance the cayenne pepper with some sweetness to avoid any stomach problems. Regular intake of this supplement keeps your body fit and active for the whole day without any tiredness. The price of the supplement is high but you can find it on discount online.
The black pepper and cayenne pepper extracts in this supplement make the body burn the excess fat at a faster rate. The veggie capsules do not have any side effects which make this product safe for women aiming for weight loss in a safe and effective manner.
Designed especially for women and marketed as a lifestyle product rather than a supplement, this fat burner from Alpha Woman is the preferred thermogenic supplement of thousands of women across the globe. It not only works on burning the fat but also controls the cortisol hormone which is responsible for all the stress and the weight gain in women. The supplement enhances the mood and boosts the natural libido in women. The formula of this product contains natural herbal ingredients like ashwagandha which are known to improve the metabolism and energy levels of the body. All these ingredients are completely safe and do not have any side effects. The supplement works on the problem areas of the body and helps to gain that hourglass figure. The price of the product is very high but the multiple benefits it has are worth it. You can buy more quantity at a lesser price if the supplement suits your body.
This supplement has much more to offer than just weight loss as it elevates mood, decreases cravings, and boosts the libido while burning fat from the body. It also controls stress which is an added advantage.
With two yummy flavors to choose from, this whey protein isolate with amino acids is the perfect powder supplement for mixing in your morning breakfast pancakes, smoothies, muffins etc. The supplement also contains grapeseed extracts, goji berry, pomegranate, cranberry, and apple which make it full of nutrients for a great post-workout meal. It tastes great and gives your body a boost of energy for accelerating the fat-burning process. It burns calories faster than other protein supplements as the thermogenic process kicks in soon after you take the supplement. It is a great option for people who do not like taking capsules as supplements. The packaging of the supplement is travel-friendly. Both the flavors are good in terms of taste so you can pick the one which you like. This is a very expensive supplement as it costs more than any other supplements on this list but the taste and the results are great.
Powder protein supplements that taste good are hard to find but this product from Shredz is just perfect. It is great in terms of taste and helps you shed those extra kilos with its punch of protein and metabolism-boosting ingredients.
Choose supplements which contain proven fat-burning supplements like raspberry ketones, Garcinia Cambogia, Green tea extracts, caffeine, whey protein, and peppers. The formula should be made up of natural ingredients and not synthetic fillers which provide instant energy and pump the body with caffeine for a very short period of time. Do not buy products which interfere with the normal female hormonal cycle of the body as these hormones perform important functions.
Read the label of the product carefully and consult your doctor to understand more about the recommended dosage of the supplement. Do not go over the daily recommended limit for any supplement as this can do more harm than good to your body. Generally, supplements are to be taken twice a day and they come in a bottle of 60 capsules which are a month's supply. Supplements lose their potency over time so do not buy them in bulk. Buy small bottles if you are a frequent traveler.
A few normal side-effects are normal for any supplement as your body takes some time to adjust to the composition but any major side-effects should be reported to a doctor. Some supplements have more harmful side effects like heart palpitations, jitters, and insomnia. Do not continue taking the supplement if you experience any of these symptoms. Look for certifications and talk to people who have used the supplements to get more information on the side-effects.
The most important thing to consider about supplements is what other users have experienced when they used the product. Reading customer reviews online will give you valuable information about the results of the supplement and whether it has any side-effects or not. You can also share your experiences of the supplement online so that others can read them and benefit from the information.
Thermogenic supplements can be a little expensive as they contain natural ingredients which are difficult to find and process. You can buy a good-quality thermogenic supplement for women in around 20$ to 50$ depending on the ingredients and the brand. Do not experiment with your body by buying cheap supplements which can damage your body and have many harmful side-effects.
Do not consume supplements without consulting your doctor in case you have a history of any disease. Many supplements are such that they can cause serious harm to life if taken by a diabetic person or a person with a heart condition.
Make sure that you combine the supplements with a right diet and exercise routine as only taking supplements will harm the body rather than benefit it. Using the capsules just before a workout improves the performance of the muscles considerably.
Stay away from supplements which contain caffeine if you are sensitive to caffeine. Such supplements can alter the heart rate and disrupt the sleep if taken at the wrong time of the day.
Stick to simple supplements made up of natural ingredients and avoid products which contain added sugars and other sweeteners which can stimulate your body for a short period of time rather than working on fat loss.
Keep your weight in control and burn fat faster by taking the supplements in the morning after your workout as this is the ideal time for such products.
Avoid formulas which contain proprietary blends which do not specify the concentration of each ingredient. Too much of any substance can harm your body and make you dependent on the formula.
Choose between different types of supplements like whey powders, capsules, dietary supplements, etc as per your liking as all these types are equally popular.
Use thermogenic supplements carefully in summers as they enhance the perspiration considerably and make you feel uncomfortable and dehydrated.
So these are five of the most popular thermogenic supplements for women which are great in terms of results. They are completely safe for consumption and have no extreme side effects. These supplements will help you reach your target body size with regular use. Choose the supplement which suits your body and is designed for the goal you are aiming for. This buyer guide will help you find the right thermogenic for your body.
I wonder if there is any effect if I take a thermogenic supplements during pregnancy?
It'd better take a thermogenic supplement before getting pregnant. It's not good for moms during pregnant. Some brands of the supplements were laced with heavy metals. If you bring all pack of supplements to consult the doctor, I'm sure that he must turn out almost of them. However, a pregnant must be caution of diet, medicine, or any other things around you. Also, you should avoid what may be worse to you and your baby.
Is it possible if I take thermogenic supplement in empty stomach?
Sure. Why not! But you have to make sure that your stomach must not be sensitive. I suggest you to have Thermogenic Supplements once after meal or at least with some snack to keep your stomach comfort.
ALPHA WOMAN Fat Burner is my favorite.


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Knight Rider Season 4 Episode 6
When a biker gang hi-jack and steal the FLAG truck, hospitalising RC3 in the process, they expect it to be full of liquor – so are amazed to find it full of computer equipment. The son of two computer experts, himself a computer genius, is persuaded into helping the gang to use the truck's advanced technology to aid them on a crime spree. Michael and K.I.T.T. must stop the gang before they use the computers to disrupt the test-run of a new computerised vehicle that is armed with a warhead…
Serie: Knight Rider
Director: Charles Bail
Guest Star: Charles Fleischer, Gary Hershberger, Julie Ronnie, Kandace Kuehl, Mark Schneider, Nancy Omi, Scott Valentine, Suzanne Rogers, Ted Petersen, William Joyce
Episode Title: The Wrong Crowd
An hour-long competition show that seeks to find the most talented and versatile body painters in the country.
Cupcake Wars
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The Fugitive (Quibi) aired on 2020 and belongs to the following categories: Drama. The IMDB rating is 10 (updated 08-19-2020). Tv-Show directed by Nick Santora. Quibi aired 1 seasons and…
Paranormal investigator Zak Bagans and his crew, Nick Groff and Aaron Goodwin, search for haunted locations both domestically and internationally. During their investigations, Zak and crew acquaint themselves with the…
Genre: Documentary, Mystery, Reality shows
Mighty Magiswords
Dae Jang Geum
Starring Lee Young Ae, it tells the tale of an orphaned kitchen cook who went on to become the king's first female physician. In a time when women held little…
Mighty Planes
Mighty Planes explores the art and science of aviation, on planes that soar to faraway places on astounding missions. From preparation, loading and take off, through the journey and to…
In a working-class neighborhood outside Los Angeles, Mike and Peggy raise eight boisterous boys. There are 10 people, three bedrooms, one bathroom and everyone in it for themselves.
Absolutely is a popular UK television comedy sketch show shown on Channel 4 between 1989 and 1993. The cast and crew were mainly Scottish; the principal writers and performers were…
Prepare for an all-new game show event where the smartest people in the country try to achieve the seemingly impossible task of answering 500 of the most difficult general knowledge…
When successful high-flier Frankie Bell is brought crashing to earth by chronic kidney failure she targets an alternate future. Eight years on she is in her second year as a…
Cha Ji-Won is a UDT officer of the Navy Special Operations Force. He is positive and has personal charm, but he falls into a desperate situation due to a friend's…


text:
A time-dependent role for the transcription factor CREB in neuronal allocation to an engram underlying a fear memory revealed using a novel in vivo optogenetic tool to modulate CREB function.
blue PYP HEK293T mouse in vivo primary mouse hippocampal neurons Endogenous gene expression
Park, A
Jacob, AD
Walters, BJ
Park, S
Rashid, AJ
Jung, JH
Lau, J
Frankland, PW
Josselyn, SA
Neuropsychopharmacology, 14 Dec 2019 DOI: 10.1038/s41386-019-0588-0 Link to full text
Abstract: The internal representation of an experience is thought to be encoded by long-lasting physical changes to the brain ("engrams") (Josselyn et al. Nat Rev Neurosci 16:521-534, 2015; Josselyn et al. J Neurosci 37:4647-4657, 2017; Schacter. 2001; Tonegawa et al. Neuron 87:918-931, 2015). Previously, we (Han et al. Science 316:457-460, 2007) and others (Zhou et al. Nat Neurosci 12:1438-1443, 2009) showed within the lateral amygdala (LA), a region critical for auditory conditioned fear, eligible neurons compete against one other for allocation to an engram. Neurons with relatively higher function of the transcription factor CREB were more likely to be allocated to the engram. In these studies, though, CREB function was artificially increased for several days before training. Precisely when increased CREB function is important for allocation remains an unanswered question. Here, we took advantage of a novel optogenetic tool (opto-DN-CREB) (Ali et al. Chem Biol 22:1531-1539, 2015) to gain spatial and temporal control of CREB function in freely behaving mice. We found increasing CREB function in a small, random population of LA principal neurons in the minutes-hours, but not 24 h, before training was sufficient to enhance memory, likely because these neurons were preferentially allocated to the underlying engram. However, similarly increasing CREB activity in a small population of random LA neurons immediately after training disrupted subsequent memory retrieval, likely by disrupting the precise spatial and temporal patterns of offline post-training neuronal activity and/or function required for consolidation. These findings reveal the importance of the timing of CREB activity in regulating allocation and subsequent memory retrieval, and further, highlight the potential of optogenetic approaches to control protein function with temporal specificity in behaving animals.
Structural Basis of Design and Engineering for Advanced Plant Optogenetics.
blue green red UV BLUF domains Cobalamin-binding domains Cryptochromes Fluorescent proteins LOV domains Phytochromes UV receptors Review
Banerjee, S
Mitra, D
Trends Plant Sci, 4 Nov 2019 DOI: 10.1016/j.tplants.2019.10.002 Link to full text
Abstract: In optogenetics, light-sensitive proteins are specifically expressed in target cells and light is used to precisely control the activity of these proteins at high spatiotemporal resolution. Optogenetics initially used naturally occurring photoreceptors to control neural circuits, but has expanded to include carefully designed and engineered photoreceptors. Several optogenetic constructs are based on plant photoreceptors, but their application to plant systems has been limited. Here, we present perspectives on the development of plant optogenetics, considering different levels of design complexity. We discuss how general principles of light-driven signal transduction can be coupled with approaches for engineering protein folding to develop novel optogenetic tools. Finally, we explore how the use of computation, networks, circular permutation, and directed evolution could enrich optogenetics.
Detection of Incorporation of p-Coumaric Acid into Photoactive Yellow Protein Variants in Vivo.
blue Fluorescent proteins Background
Brechun, KE
Zhen, D
Borisenko, V
Kumauchi, M
Hoff, WD
Arndt, KM
Biochemistry, 30 May 2019 DOI: 10.1021/acs.biochem.9b00279 Link to full text
Abstract: We report the design and characterization of photoactive yellow protein (PYP)-blue fluorescent protein (mTagBFP) fusion constructs that permit the direct assay of reconstitution and function of the PYP domain. These constructs allow for in vivo testing of co-expression systems for enzymatic production of the p-coumaric acid-based PYP chromophore, via the action of tyrosine ammonia lyase and p-coumaroyl-CoA ligase (pCL or 4CL). We find that different 4CL enzymes can function to reconstitute PYP, including 4CL from Arabidopsis thaliana that can produce ∼100% holo-PYP protein under optimal conditions. mTagBFP fusion constructs additionally enable rapid analysis of effects of mutations on PYP photocycles. We use this mTagBFP fusion strategy to demonstrate in vivo reconstitution of several PYP-based optogenetic tools in Escherichia coli via a biosynthesized chromophore, an important step for the use of these optogenetic tools in vivo in diverse hosts.
A yeast system for discovering optogenetic inhibitors of eukaryotic translation initiation.
blue cyan AsLOV2 Dronpa145K/N PYP RsLOV S. cerevisiae
Lu, H
Mazumder, M
Jaikaran, ASI
Kumar, A
Leis, E
Altmann, M
Cochrane, A
ACS Synth Biol, 22 Mar 2019 DOI: 10.1021/acssynbio.8b00386 Link to full text
Abstract: The precise spatiotemporal regulation of protein synthesis is essential for many complex biological processes such as memory formation, embryonic development and tumor formation. Current methods used to study protein synthesis offer only a limited degree of spatiotemporal control. Optogenetic methods, in contrast, offer the prospect of controlling protein synthesis non-invasively within minutes and with a spatial scale as small as a single synapse. Here, we present a hybrid yeast system where growth depends on the activity of human eukaryotic initiation factor 4E (eIF4E) that is suitable for screening optogenetic designs for the down-regulation of protein synthesis. We used this system to screen a diverse initial panel of 15 constructs designed to couple a light switchable domain (PYP, RsLOV, LOV, Dronpa) to 4EBP2 (eukaryotic initiation factor 4E binding protein 2), a native inhibitor of translation initiation. We identified cLIPS1 (circularly permuted LOV inhibitor of protein synthesis 1), a fusion of a segment of 4EBP2 and a circularly permuted version of the LOV2 domain from Avena sativa, as a photo-activated inhibitor of translation. Adapting the screen for higher throughput, we tested small libraries of cLIPS1 variants and found cLIPS2, a construct with an improved degree of optical control. We show that these constructs can both inhibit translation in yeast harboring a human eIF4E in vivo, and bind human eIF4E in vitro in a light-dependent manner. This hybrid yeast system thus provides a convenient way for discovering optogenetic constructs that can regulate of human eIF4E-depednednt translation initiation in a mechanistically defined manner.
Blue-Light Receptors for Optogenetics.
blue green red UV BLUF domains Cryptochromes Fluorescent proteins LOV domains Opsins Phytochromes UV receptors Review
Losi, A
Gardner, KH
Möglich, A
Chem Rev, 9 Jul 2018 DOI: 10.1021/acs.chemrev.8b00163 Link to full text
Abstract: Sensory photoreceptors underpin light-dependent adaptations of organismal physiology, development, and behavior in nature. Adapted for optogenetics, sensory photoreceptors become genetically encoded actuators and reporters to enable the noninvasive, spatiotemporally accurate and reversible control by light of cellular processes. Rooted in a mechanistic understanding of natural photoreceptors, artificial photoreceptors with customized light-gated function have been engineered that greatly expand the scope of optogenetics beyond the original application of light-controlled ion flow. As we survey presently, UV/blue-light-sensitive photoreceptors have particularly allowed optogenetics to transcend its initial neuroscience applications by unlocking numerous additional cellular processes and parameters for optogenetic intervention, including gene expression, DNA recombination, subcellular localization, cytoskeleton dynamics, intracellular protein stability, signal transduction cascades, apoptosis, and enzyme activity. The engineering of novel photoreceptors benefits from powerful and reusable design strategies, most importantly light-dependent protein association and (un)folding reactions. Additionally, modified versions of these same sensory photoreceptors serve as fluorescent proteins and generators of singlet oxygen, thereby further enriching the optogenetic toolkit. The available and upcoming UV/blue-light-sensitive actuators and reporters enable the detailed and quantitative interrogation of cellular signal networks and processes in increasingly more precise and illuminating manners.
Optogenetics: A Primer for Chemists.
blue green near-infrared red UV BLUF domains Cobalamin-binding domains Cryptochromes Fluorescent proteins LOV domains Phytochromes UV receptors Review
Lawrence, DS
O'Banion, CP
Chembiochem, 19 Apr 2018 DOI: 10.1002/cbic.201800013 Link to full text
Abstract: The field of optogenetics uses genetically encoded, light-responsive proteins to control physiological processes. This technology has been hailed as the one of the ten big ideas in brain science in the past decade,[1] the breakthrough of the decade,[2] and the method of the year in 2010[3] and again in 2014[4]. The excitement evidenced by these proclamations is confirmed by a couple of impressive numbers. The term "optogenetics" was coined in 2006.[5] As of December 2017, "optogenetics" is found in the title or abstract of almost 1600 currently funded National Institutes of Health grants. In addition, nearly 600 reviews on optogenetics have appeared since 2006, which averages out to approximately one review per week! However, in spite of these impressive numbers, the potential applications and implications of optogenetics are not even close to being fully realized. This is due, in large part, to the challenges associated with the design of optogenetic analogs of endogenous proteins. This review is written from a chemist's perspective, with a focus on the molecular strategies that have been developed for the construction of optogenetic proteins.
Strategies for the photo-control of endogenous protein activity.
blue Cryptochromes Fluorescent proteins LOV domains Review
Curr Opin Struct Biol, 28 Nov 2016 DOI: 10.1016/j.sbi.2016.11.014 Link to full text
Abstract: Photo-controlled or 'optogenetic' effectors interfacing with endogenous protein machinery allow the roles of endogenous proteins to be probed. There are two main approaches being used to develop optogenetic effectors: (i) caging strategies using photo-controlled conformational changes, and (ii) protein relocalization strategies using photo-controlled protein-protein interactions. Numerous specific examples of these approaches have been reported and efforts to develop general methods for photo-control of endogenous proteins are a current focus. The development of improved screening and selection methods for photo-switchable proteins would advance the field.
Natural Resources for Optogenetic Tools.
blue green red UV Cobalamin-binding domains Cryptochromes Fluorescent proteins LOV domains Opsins Phytochromes UV receptors Review
Mathes, T
Methods Mol Biol, 2016 DOI: 10.1007/978-1-4939-3512-3_2 Link to full text
Abstract: Photoreceptors are found in all kingdoms of life and mediate crucial responses to environmental challenges. Nature has evolved various types of photoresponsive protein structures with different chromophores and signaling concepts for their given purpose. The abundance of these signaling proteins as found nowadays by (meta-)genomic screens enriched the palette of optogenetic tools significantly. In addition, molecular insights into signal transduction mechanisms and design principles from biophysical studies and from structural and mechanistic comparison of homologous proteins opened seemingly unlimited possibilities for customizing the naturally occurring proteins for a given optogenetic task. Here, a brief overview on the photoreceptor concepts already established as optogenetic tools in natural or engineered form, their photochemistry and their signaling/design principles is given. Finally, so far not regarded photosensitive modules and protein architectures with potential for optogenetic application are described.
Optogenetic Inhibitor of the Transcription Factor CREB.
blue PYP HEK293T in vitro primary mouse cortical neurons Endogenous gene expression
Ali, AM
Xia, Y
Mercaldo, V
Karanicolas, J
Chem Biol, 19 Nov 2015 DOI: 10.1016/j.chembiol.2015.09.018 Link to full text
Abstract: Current approaches for optogenetic control of transcription do not mimic the activity of endogenous transcription factors, which act at numerous sites in the genome in a complex interplay with other factors. Optogenetic control of dominant negative versions of endogenous transcription factors provides a mechanism for mimicking the natural regulation of gene expression. Here we describe opto-DN-CREB, a blue-light-controlled inhibitor of the transcription factor CREB created by fusing the dominant negative inhibitor A-CREB to photoactive yellow protein (PYP). A light-driven conformational change in PYP prevents coiled-coil formation between A-CREB and CREB, thereby activating CREB. Optogenetic control of CREB function was characterized in vitro, in HEK293T cells, and in neurons where blue light enabled control of expression of the CREB targets NR4A2 and c-Fos. Dominant negative inhibitors exist for numerous transcription factors; linking these to optogenetic domains offers a general approach for spatiotemporal control of native transcriptional events.
Applications of hydrogen deuterium exchange (HDX) for the characterization of conformational dynamics in light-activated photoreceptors.
blue red UV BLUF domains Fluorescent proteins LOV domains Phytochromes UV receptors Review
Lindner, R
Heintz, U
Winkler, A
Front Mol Biosci, 23 Jun 2015 DOI: 10.3389/fmolb.2015.00033 Link to full text
Abstract: Rational design of optogenetic tools is inherently linked to the understanding of photoreceptor function. Structural analysis of elements involved in signal integration in individual sensor domains provides an initial idea of their mode of operation, but understanding how local structural rearrangements eventually affect signal transmission to output domains requires inclusion of the effector regions in the characterization. However, the dynamic nature of these assemblies renders their structural analysis challenging and therefore a combination of high- and low-resolution techniques is required to appreciate functional aspects of photoreceptors. This review focuses on the potential of hydrogen-deuterium exchange coupled to mass spectrometry (HDX-MS) for complementing the structural characterization of photoreceptors. In this respect, the ability of HDX-MS to provide information on conformational dynamics and the possibility to address multiple functionally relevant states in solution render this methodology ideally suitable. We highlight recent examples demonstrating the potential of HDX-MS and discuss how these results can help to improve existing optogenetic systems or guide the design of novel optogenetic tools.
Time-resolved crystallography and protein design: signalling photoreceptors and optogenetics.
blue Fluorescent proteins Review
Moffat, K
Philos Trans R Soc Lond, B, Biol Sci, 17 Jul 2014 DOI: 10.1098/rstb.2013.0568 Link to full text
Abstract: Time-resolved X-ray crystallography and solution scattering have been successfully conducted on proteins on time-scales down to around 100 ps, set by the duration of the hard X-ray pulses emitted by synchrotron sources. The advent of hard X-ray free-electron lasers (FELs), which emit extremely intense, very brief, coherent X-ray pulses, opens the exciting possibility of time-resolved experiments with femtosecond time resolution on macromolecular structure, in both single crystals and solution. The X-ray pulses emitted by an FEL differ greatly in many properties from those emitted by a synchrotron, in ways that at first glance make time-resolved measurements of X-ray scattering with the required accuracy extremely challenging. This opens up several questions which I consider in this brief overview. Are there likely to be chemically and biologically interesting structural changes to be revealed on the femtosecond time-scale? How shall time-resolved experiments best be designed and conducted to exploit the properties of FELs and overcome challenges that they pose? To date, fast time-resolved reactions have been initiated by a brief laser pulse, which obviously requires that the system under study be light-sensitive. Although this is true for proteins of the visual system and for signalling photoreceptors, it is not naturally the case for most interesting biological systems. To generate more biological targets for time-resolved study, can this limitation be overcome by optogenetic, chemical or other means?
Optical control of protein-protein interactions via blue light-induced domain swapping.
Burns, DC
Biochemistry, 16 Jul 2014 DOI: 10.1021/bi500622x Link to full text
Abstract: The design of new optogenetic tools for controlling protein function would be facilitated by the development of protein scaffolds that undergo large, well-defined structural changes upon exposure to light. Domain swapping, a process in which a structural element of a monomeric protein is replaced by the same element of another copy of the same protein, leads to a well-defined change in protein structure. We observe domain swapping in a variant of the blue light photoreceptor photoactive yellow protein in which a surface loop is replaced by a well-characterized protein-protein interaction motif, the E-helix. In the domain-swapped dimer, the E-helix sequence specifically binds a partner K-helix sequence, whereas in the monomeric form of the protein, the E-helix sequence is unable to fold into a binding-competent conformation and no interaction with the K-helix is seen. Blue light irradiation decreases the extent of domain swapping (from Kd = 10 μM to Kd = 300 μM) and dramatically enhances the rate, from weeks to <1 min. Blue light-induced domain swapping thus provides a novel mechanism for controlling of protein-protein interactions in which light alters both the stability and the kinetic accessibility of binding-competent states.
Proteins in action: femtosecond to millisecond structural dynamics of a photoactive flavoprotein.
Brust, R
Lukacs, A
Haigney, A
Addison, K
Gil, AA
Towrie, M
Clark, IP
Greetham, GM
Tonge, PJ
Meech, SR
J Am Chem Soc, 22 Oct 2013 DOI: 10.1021/ja407265p Link to full text
Abstract: Living systems are fundamentally dependent on the ability of proteins to respond to external stimuli. The mechanism, the underlying structural dynamics, and the time scales for regulation of this response are central questions in biochemistry. Here we probe the structural dynamics of the BLUF domain found in several photoactive flavoproteins, which is responsible for light activated functions as diverse as phototaxis and gene regulation. Measurements have been made over 10 decades of time (from 100 fs to 1 ms) using transient vibrational spectroscopy. Chromophore (flavin ring) localized dynamics occur on the pico- to nanosecond time scale, while subsequent protein structural reorganization is observed over microseconds. Multiple time scales are observed for the dynamics associated with different vibrations of the protein, suggesting an underlying hierarchical relaxation pathway. Structural evolution in residues directly H-bonded to the chromophore takes place more slowly than changes in more remote residues. However, a point mutation which suppresses biological function is shown to 'short circuit' this structural relaxation pathway, suppressing the changes which occur further away from the chromophore while accelerating dynamics close to it.
A circularly permuted photoactive yellow protein as a scaffold for photoswitch design.
Al-Abdul-Wahid, MS
Biochemistry, 1 May 2013 DOI: 10.1021/bi400018h Link to full text
Abstract: Upon blue light irradiation, photoactive yellow protein (PYP) undergoes a conformational change that involves large movements at the N-terminus of the protein. We reasoned that this conformational change might be used to control other protein or peptide sequences if these were introduced as linkers connecting the N- and C-termini of PYP in a circular permutant. For such a design strategy to succeed, the circularly permuted PYP (cPYP) would have to fold normally and undergo a photocycle similar to that of the wild-type protein. We created a test cPYP by connecting the N- and C-termini of wild-type PYP (wtPYP) with a GGSGGSGG linker polypeptide and introducing new N- and C-termini at G115 and S114, respectively. Biophysical analysis indicated that this cPYP adopts a dark-state conformation much like wtPYP and undergoes wtPYP-like photoisomerization driven by blue light. However, thermal recovery of dark-state cPYP is ∼10-fold faster than that of wtPYP, so that very bright light is required to significantly populate the light state. Targeted mutations at M121E (M100 in wtPYP numbering) were found to enhance the light sensitivity substantially by lengthening the lifetime of the light state to ∼10 min. Nuclear magnetic resonance (NMR), circular dichroism, and UV-vis analysis indicated that the M121E-cPYP mutant also adopts a dark-state structure like that of wtPYP, although protonated and deprotonated forms of the chromophore coexist, giving rise to a shoulder near 380 nm in the UV-vis absorption spectrum. Fluorine NMR studies with fluorotryptophan-labeled M121E-cPYP show that blue light drives large changes in conformational dynamics and leads to solvent exposure of Trp7 (Trp119 in wtPYP numbering), consistent with substantial rearrangement of the N-terminal cap structure. M121E-cPYP thus provides a scaffold that may allow a wider range of photoswitchable protein designs via replacement of the linker polypeptide with a target protein or peptide sequence.
Genetically engineered light sensors for control of bacterial gene expression.
blue green red Cyanobacteriochromes Fluorescent proteins LOV domains Phytochromes Review
Camsund, D
Lindblad, P
Jaramillo, A
Biotechnol J, 7 Jun 2011 DOI: 10.1002/biot.201100091 Link to full text
Abstract: Light of different wavelengths can serve as a transient, noninvasive means of regulating gene expression for biotechnological purposes. Implementation of advanced gene regulatory circuits will require orthogonal transcriptional systems that can be simultaneously controlled and that can produce several different control states. Fully genetically encoded light sensors take advantage of the favorable characteristics of light, do not need the supplementation of any chemical inducers or co-factors, and have been demonstrated to control gene expression in Escherichia coli. Herein, we review engineered light-sensor systems with potential for in vivo regulation of gene expression in bacteria, and highlight different means of extending the range of available light input and transcriptional output signals. Furthermore, we discuss advances in multiplexing different light sensors for achieving multichromatic control of gene expression and indicate developments that could facilitate the construction of efficient systems for light-regulated, multistate control of gene expression.
The short-lived signaling state of the photoactive yellow protein photoreceptor revealed by combined structural probes.
Ramachandran, PL
Lovett, JE
Carl, PJ
Cammarata, M
Lee, JH
Jung, YO
Ihee, H
Timmel, CR
van Thor, JJ
J Am Chem Soc, 31 Mar 2011 DOI: 10.1021/ja200617t Link to full text
Abstract: The signaling state of the photoactive yellow protein (PYP) photoreceptor is transiently developed via isomerization of its blue-light-absorbing chromophore. The associated structural rearrangements have large amplitude but, due to its transient nature and chemical exchange reactions that complicate NMR detection, its accurate three-dimensional structure in solution has been elusive. Here we report on direct structural observation of the transient signaling state by combining double electron electron resonance spectroscopy (DEER), NMR, and time-resolved pump-probe X-ray solution scattering (TR-SAXS/WAXS). Measurement of distance distributions for doubly spin-labeled photoreceptor constructs using DEER spectroscopy suggests that the signaling state is well ordered and shows that interspin-label distances change reversibly up to 19 Å upon illumination. The SAXS/WAXS difference signal for the signaling state relative to the ground state indicates the transient formation of an ordered and rearranged conformation, which has an increased radius of gyration, an increased maximum dimension, and a reduced excluded volume. Dynamical annealing calculations using the DEER derived long-range distance restraints in combination with short-range distance information from (1)H-(15)N HSQC perturbation spectroscopy give strong indication for a rearrangement that places part of the N-terminal domain in contact with the exposed chromophore binding cleft while the terminal residues extend away from the core. Time-resolved global structural information from pump-probe TR-SAXS/WAXS data supports this conformation and allows subsequent structural refinement that includes the combined energy terms from DEER, NMR, and SAXS/WAXS together. The resulting ensemble simultaneously satisfies all restraints, and the inclusion of TR-SAXS/WAXS effectively reduces the uncertainty arising from the possible spin-label orientations. The observations are essentially compatible with reduced folding of the I(2)' state (also referred to as the 'pB' state) that is widely reported, but indicates it to be relatively ordered and rearranged. Furthermore, there is direct evidence for the repositioning of the N-terminal region in the I(2)' state, which is structurally modeled by dynamical annealing and refinement calculations.
Lights on and action! Controlling microbial gene expression by light.
blue green near-infrared red BLUF domains Cryptochromes Cyanobacteriochromes Fluorescent proteins LOV domains Phytochromes Review
Drepper, T
Krauss, U
Meyer zu Berstenhorst, S
Pietruszka, J
Jaeger, KE
Appl Microbiol Biotechnol, 20 Feb 2011 DOI: 10.1007/s00253-011-3141-6 Link to full text
Abstract: Light-mediated control of gene expression and thus of any protein function and metabolic process in living microbes is a rapidly developing field of research in the areas of functional genomics, systems biology, and biotechnology. The unique physical properties of the environmental factor light allow for an independent photocontrol of various microbial processes in a noninvasive and spatiotemporal fashion. This mini review describes recently developed strategies to generate photo-sensitive expression systems in bacteria and yeast. Naturally occurring and artificial photoswitches consisting of light-sensitive input domains derived from different photoreceptors and regulatory output domains are presented and individual properties of light-controlled expression systems are discussed.
Tripping the light fantastic: blue-light photoreceptors as examples of environmentally modulated protein-protein interactions.
Zoltowski, BD
Biochemistry, 14 Dec 2010 DOI: 10.1021/bi101665s Link to full text
Abstract: Blue-light photoreceptors play a pivotal role in detecting the quality and quantity of light in the environment, controlling a wide range of biological responses. Several families of blue-light photoreceptors have been characterized in detail using biophysics and biochemistry, beginning with photon absorption, through intervening signal transduction, to regulation of biological activities. Here we review the light oxygen voltage, cryptochrome, and sensors of blue light using FAD families, three different groups of proteins that offer distinctly different modes of photochemical activation and signal transduction yet play similar roles in a vast array of biological responses. We cover mechanisms of light activation and propagation of conformational responses that modulate protein-protein interactions involved in biological signaling. Discovery and characterization of these processes in natural proteins are now allowing the design of photoregulatable engineered proteins, facilitating the generation of novel reagents for biochemical and cell biological research.
A photoswitchable DNA-binding protein based on a truncated GCN4-photoactive yellow protein chimera.
blue PYP in vitro
Morgan, SA
Photochem Photobiol Sci, 13 Sep 2010 DOI: 10.1039/c0pp00214c Link to full text
Abstract: Photo-controlled DNA-binding proteins promise to be useful tools for probing complex spatiotemporal patterns of gene expression in living organisms. Here we report a novel photoswitchable DNA-binding protein, GCN4(S)Δ25PYP, based on a truncated GCN4-photoactive yellow protein chimera. In contrast to previously reported designed photoswitchable proteins where DNA binding affinity is enhanced upon irradiation, GCN4(S)Δ25PYP dissociates from DNA when irradiated with blue light. In addition, the rate of thermal relaxation to the ground state, part of the PYP photocycle, is enhanced by DNA binding whereas in previous reported constructs it is slowed. The origins of this reversed photoactivity are analyzed in structural terms.
Structure and function of plant photoreceptors.
blue near-infrared red Cryptochromes Fluorescent proteins LOV domains Phytochromes Review Background
Yang, X
Ayers, RA
Annu Rev Plant Biol, 25 Jan 2010 DOI: 10.1146/annurev-arplant-042809-112259 Link to full text
Abstract: Signaling photoreceptors use the information contained in the absorption of a photon to modulate biological activity in plants and a wide range of organisms. The fundamental-and as yet imperfectly answered-question is, how is this achieved at the molecular level? We adopt the perspective of biophysicists interested in light-dependent signal transduction in nature and the three-dimensional structures that underpin signaling. Six classes of photoreceptors are known: light-oxygen-voltage (LOV) sensors, xanthopsins, phytochromes, blue-light sensors using flavin adenine dinucleotide (BLUF), cryptochromes, and rhodopsins. All are water-soluble proteins except rhodopsins, which are integral membrane proteins; all are based on a modular architecture except cryptochromes and rhodopsins; and each displays a distinct, light-dependent chemical process based on the photochemistry of their nonprotein chromophore, such as isomerization about a double bond (xanthopsins, phytochromes, and rhodopsins), formation or rupture of a covalent bond (LOV sensors), or electron transfer (BLUF sensors and cryptochromes).
Photoregulation in prokaryotes.
blue near-infrared red Fluorescent proteins LOV domains Phytochromes Review Background
Purcell, EB
Crosson, S
Curr Opin Microbiol, 8 Apr 2008 DOI: 10.1016/j.mib.2008.02.014 Link to full text
Abstract: The spectroscopic identification of sensory rhodopsin I by Bogomolni and Spudich in 1982 provided a molecular link between the light environment and phototaxis in Halobacterium salinarum, and thus laid the foundation for the study of signal transducing photosensors in prokaryotes. In recent years, a number of new prokaryotic photosensory receptors have been discovered across a broad range of taxa, including dozens in chemotrophic species. Among these photoreceptors are new classes of rhodopsins, BLUF-domain proteins, bacteriophytochromes, cryptochromes, and LOV-family photosensors. Genetic and biochemical analyses of these receptors have demonstrated that they can regulate processes ranging from photosynthetic pigment biosynthesis to virulence.
Structure and photoreaction of photoactive yellow protein, a structural prototype of the PAS domain superfamily.
Imamoto, Y
Kataoka, M
Photochem Photobiol, 1 Jan 2007 DOI: 10.1562/2006-02-28-ir-827 Link to full text
Abstract: Photoactive yellow protein (PYP) is a water-soluble photosensor protein found in purple photosynthetic bacteria. Unlike bacterial rhodopsins, photosensor proteins composed of seven transmembrane helices and a retinal chromophore in halophilic archaebacteria, PYP is a highly soluble globular protein. The alpha/beta fold structure of PYP is a structural prototype of the PAS domain superfamily, many members of which function as sensors for various kinds of stimuli. To absorb a photon in the visible region, PYP has a p-coumaric acid chromophore binding to the cysteine residue via a thioester bond. It exists in a deprotonated trans form in the dark. The primary photochemical event is photo-isomerization of the chromophore from trans to cis form. The twisted cis chromophore in early intermediates is relaxed and finally protonated. Consequently, the chromophore becomes electrostatically neutral and rearrangement of the hydrogen-bonding network triggers overall structural change of the protein moiety, in which local conformational change around the chromophore is propagated to the N-terminal region. Thus, it is an ideal model for protein conformational changes that result in functional change, responding to stimuli and expressing physiological activity. In this paper, recent progress in investigation of the photoresponse of PYP is reviewed.
The LOV domain family: photoresponsive signaling modules coupled to diverse output domains.
blue Fluorescent proteins LOV domains Review Background
Rajagopal, S
Biochemistry, 14 Jan 2003 DOI: 10.1021/bi026978l Link to full text
Abstract: For single-cell and multicellular systems to survive, they must accurately sense and respond to their cellular and extracellular environment. Light is a nearly ubiquitous environmental factor, and many species have evolved the capability to respond to this extracellular stimulus. Numerous photoreceptors underlie the activation of light-sensitive signal transduction cascades controlling these responses. Here, we review the properties of the light, oxygen, or voltage (LOV) family of blue-light photoreceptor domains, a subset of the Per-ARNT-Sim (PAS) superfamily. These flavin-binding domains, first identified in the higher-plant phototropins, are now shown to be present in plants, fungi, and bacteria. Notably, LOV domains are coupled to a wide array of other domains, including kinases, phosphodiesterases, F-box domains, STAS domains, and zinc fingers, which suggests that the absorption of blue light by LOV domains regulates the activity of these structurally and functionally diverse domains. LOV domains contain a conserved molecular volume extending from the flavin cofactor, which is the locus for light-driven structural change, to the molecular surface. We discuss the role of this conserved volume of structure in LOV-regulated processes.
BLUF: a novel FAD-binding domain involved in sensory transduction in microorganisms.
blue red BLUF domains Fluorescent proteins LOV domains Phytochromes Background
Gomelsky, M
Klug, G
Trends Biochem Sci, 1 Oct 2002 DOI: 10.1016/s0968-0004(02)02181-3 Link to full text
Abstract: A novel FAD-binding domain, BLUF, exemplified by the N-terminus of the AppA protein from Rhodobacter sphaeroides, is present in various proteins, primarily from Bacteria. The BLUF domain is involved in sensing blue-light (and possibly redox) using FAD and is similar to the flavin-binding PAS domains and cryptochromes. The predicted secondary structure reveals that the BLUF domain is a novel FAD-binding fold.


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Prey abundance and habitat during the breeding season for Piping Plovers in the Ontario Great Lakes region
Similar to other shorebird trends around the world, the Piping Plover population (Charadrius melodus circumcinctus) is projected to decline if concerted conservation efforts are relaxed. To date, there is insufficient understanding of the connection between habitat type, prey abundance, and chick behaviour of the Piping Plover breeding population in Ontario. The aim of my thesis was to gain knowledge about prey abundance at recent and historic breeding locations, understanding how habitat influences prey abundance and chick behaviour across the Piping Plover breeding range in Ontario's Great Lakes. The objective of my first study was to understand prey abundance across the breeding region Great Lakes of Ontario from 2018-2019, including occupied and unoccupied sites, and to quantify variation among habitats and periods of reproduction. To evaluate resources, I sampled 17 locations to compare prey abundance using invertebrate traps (n= 3,507). Sampling took place over the reproductive periods of nest initiation, post-hatch, and fledging and in four habitat types of shoreline, wrack, berm, and back dune. Occupied breeding sites had higher prey abundances, and different assemblages of invertebrate prey than unoccupied sites. Additionally, breeding sites had higher prey abundance during nest initiation and supported higher amounts of prey in shoreline and wrack habitat. The objective of my second study was to understand how habitat types influence chick behaviour. To evaluate behaviour-habitat trends, instantaneous chick observations were recorded at the four nest sites from the post-hatch to fledging stages. In total there were 23 fledged chicks that we observed across the two years. Chicks in this study spent 60.9% of their time foraging, 11.9% of the time displaying alert behaviour, 21.4% of their time resting or being brooded, and 5.9% of their time preening. Chicks spent a large proportion of time foraging in the shoreline, resting in the back dune, and alert in berm habitat. The frequency of these alert, defensive behaviours differed among sites, with Sauble Beach chicks spending more time in defensive behaviours compared to other sites. I concluded that in both nesting and brood-rearing periods, habitat is selected non-randomly by adult and young Piping Plovers to maximize access to invertebrate prey for growth and survival. Access by chicks to the most productive habitats should be considered in local management decisions. Author Keywords: chick behaviour, endangered, Great Lakes Region, habitat, Piping Plovers, prey abundance
Population Genetics and Gut Microbiome Composition Reveal Subdivisions and Space Use in a Generalist and Specialist Ungulate
Natural populations are often difficult and costly to study, due to the plethora of confounding processes and variables present. This is of particular importance when dealing with managed species. Ungulates, for example, act as both consumers and prey sources; they also provide economic benefit through harvest, and as such, are of high ecological and economic value. I addressed conservation and management concerns by quantifying subdivision in wild populations and combined movement with non-invasive sampling to provide novel insight on the physiological drivers of space use in multiple species. This thesis explored biological patterns in ungulates using two distinct approaches: the first used molecular genetics to quantify gene flow, while the second examined the relationship between movement and the gut microbiome using high-throughput sequencing and GPS tracking. The goal of the first chapter was to quantify gene flow and assess the population structure of mountain goats (Oreamnos americanus) in northern British Columbia (BC) to inform management. I used microsatellites to generate genotype data and used a landscape genetics framework to evaluate the possible drivers behind genetic differentiation. The same analyses were performed at both a broad and fine scale, assessing genetic differentiation between populations in all of northern BC and in a case management study area northeast of Smithers BC. The results indicated panmixia among mountain goats regardless of scale, suggesting distance and landscape resistance were minimally inhibiting gene flow. Therefore, management at local scales can continue with little need for genetically informed boundaries, but regulations should be tailored to specific regions incorporating data on local access and harvest pressure. My second chapter aimed to determine the extent to which the gut microbiome drives space-use patterns in a specialist (mountain goat) and generalist (white-tailed deer, Odocoileus virginianus) ungulate. Using fecal samples, we generated genomic data using 16S rRNA high-throughput sequencing to evaluate gut diversity and gut microbiome characteristics. Additionally, individuals were fitted with GPS collars so that we could gain insight into movement patterns. Gut microbiome metrics were stronger predictors of space use and movement patterns with respect to home range size, whereas they were weaker predictors of habitat use. Notably, factors of both the gut microbiome and age of a given species were correlated with changes in space use and habitat use. Ultimately, this research linked high-throughput sequencing and GPS data to better understand ecological processes in wild ungulates. Author Keywords: gene flow, genomics, gut microbiome, home range, population genetic structure, ungulates
Diversity, Biogeography, and Functional Traits of Native Bees from Ontario's Far North and Akimiski Island, Nunavut
Bees (clade Anthophila), are poorly studied in northern Canada, as these regions can be difficult to access and have a short growing season. This study examined bees from two such regions: Ontario's Far North, and Akimiski Island, Nunavut. I present this study as the largest biogeographical study of bees performed in these remote areas to enhance knowledge of northern native bees. I found 10 geographically unexpected species in Ontario and on Akimiski Island. Rarefaction and the Chao 1 Diversity Index showed that Akimiski is nearly as diverse as the Far North of Ontario, a significantly larger area. I also found, based on log femur length versus latitude, Bombus worker size was consistent with Bergmann's rule, and there were no apparent statistical differences in the community weighted means of functional traits between the Far North's Boreal Shield and Hudson Bay Lowlands ecozones. This work provides invaluable knowledge of the native bee species from these regions, which has implications for their future conservation. Author Keywords: Akimiski Island, Bergmann's rule, Chao 1, Community-weighted means, native bees, rarefaction
Assessment of Potential Threats to Eastern Flowering Dogwood (Cornus florida) in Southern Ontario
In Canada, eastern flowering dogwood (Cornus florida L.) is an endangered tree that occurs only in the Carolinian forest of southern Ontario. Threats to this species include habitat fragmentation and the fungal pathogen dogwood anthracnose (Discula destructiva). I conducted a population genetic analysis using seven nuclear microsatellite markers to determine if fragmented populations are genetically isolated from one another and have low levels of genetic diversity. Genetic comparisons suggest on-going dispersal among sites and relatively high genetic diversity within most sites; however, smaller populations and younger trees were less genetically diverse. I also used linear mixed effects models to assess potential relationships between several ecological variables and the prevalence of dogwood anthracnose. Disease severity was higher in trees on shallow slopes and in larger trees; the latter also had higher likelihood of infection. Insights from this study will be important to incorporate into future management strategies. Author Keywords: Cornus florida, Discula destructiva, dogwood anthracnose, Eastern flowering dogwood, endangered, population genetics
Environmental structure, morphology and spatial ecology of the five-lined skink (Plestiodon fasciatus) at high latitude range limits
Detecting relevant and meaningful patterns from the complex, interconnected network of relationships between organisms and their environment is a primary objective of ecology. Ecological patterns occur across multiple scales of space and time. In this dissertation, I examine aspects of environmental structure that influence a species' distribution and are expressed in that species' population dynamics. I compare the morphology of the five-lined skink (Plestiodon fasciatus) near its high latitude range limits with a lower latitude population and evaluate the economics of their behaviour in the context of its reproductive strategy. I tested the conformity of this species to biogeographical rules postulated by MacArthur, Bergmann, and Rensch. Spatial ecology was investigated in the context of the environmental potential for polygamy proposed by Emlen and Oring (1977) The five-lined skink, Plestiodon fasciatus, conformed to these biogeographic rules. Specifically, abiotic factors were the primary limiting factors affecting distribution at the high latitude range limits of the species; body size was larger in high latitude populations; and the degree of sexual size dimorphism was greater at high latitude than at low latitude. Spatial ecology at the individual scale was influenced by sites with suitable thermal conditions which facilitate the polygynandrous mating system documented in P. fasciatus in high latitude populations. My results confirm the importance of microsites with suitable thermal profiles as key habitat for ectothermic vertebrates at high latitudes. The influence of temperature as a limiting abiotic factor is expressed in population density, body size, spatial ecology, and reproductive strategy of P. fasciatus. Conservation and restoration of high latitude populations of ectothermic vertebrates should focus on ensuring thermal requirements of the species of concern are met before other factors are addressed, as temperature is likely the single most important limiting factor at high latitude range limits. Author Keywords: biogeography, lizard, Plestiodon fasciatus, range limits, sexual size dimorphism, spatial ecology
multi-faceted approach to evaluating the detection probability of an elusive snake (Sistrurus catenatus)
Many rare and elusive species have low detection probabilities, thereby imposing unique challenges to monitoring and conservation. Here, we assess the detection probability of the Eastern Massasauga (Sistrurus catenatus) in contrast to a more common and conspicuous species, the Eastern Gartersnake (Thamnophis sirtalis). We found that patterns of detection probability differed between species, wherein S. catenatus was detected less often and under a more specific set of sampling conditions. Correspondingly, detection trials with S. catenatus found a high non-detection rate, while detection trials with artificial models suggest that regional differences in detection probability are driven by variation in population density and habitat use. Our results suggest that current monitoring efforts are not sufficient, and that S. catenatus is frequently undetected. Accordingly, we highlight the importance of species-specific monitoring protocols when monitoring rare and elusive species, and recommend a multi-faceted approach that estimates detection probability and identifies species-specific challenges to monitoring. Author Keywords: detection probability, elusive species, monitoring programs, non-detection, S. catenatus, snakes
Effects of Local, Landscape, and Temporal Variables on Bobolink Nest Survival in Southern Ontario
Populations of grassland birds, including the Bobolink (Dolichonyx oryzivorus), are experiencing steep declines due to losses of breeding habitat, land use changes, and agricultural practices. Understanding the variables affecting reproductive success can aid conservation of grassland species. I investigated 1) whether artificial nest experiments accurately estimate the impacts of cattle on the daily survival rate of Bobolink nests and 2) which local, landscape, and temporal variables affect daily survival rate of Bobolink nests in Southern Ontario. I replicated an artificial nest experiment performed in 2012 and 2015 to compare the daily survival rate of artificial and natural nests at multiple stocking rates (number of cattle × days × ha-1). I also monitored Bobolink nests and modeled daily survival rate using local variables (e.g., stocking rate, field use, patch area), landscape variables (e.g., percent forest within 2, 5, and 10 km), and temporal variables (e.g., year, date of season). Results indicate that artificial nest experiments using clay shooting targets overestimated the impacts of stocking rate on the daily survival rate of Bobolink nests. With natural nests, region (confounded by year and field use), stocking rate, and date of season were the strongest predictors of daily survival rate; with stocking rate and date of season both having a negative effect. Management should focus on conserving pastures with low stocking rates (< 40 cattle × days × ha-1), late-cut hayfields, fallow fields, and other grasslands to protect breeding grounds for the Bobolink and other declining grassland bird species. Author Keywords: Bobolink, Daily survival rate, landscape variables, local variables, Nest survival, temporal variables
Risk of Mortality for the Semipalmated Plover (Charadrius semipalmatus) Throughout Its Life Cycle
Three long-term mark and recapture/resight data sets of individually marked Semipalmated Plovers (Charadrius semipalmatus) were analyzed using Cormack-Jolly- Seber models. Data came from two breeding populations (Churchill, Manitoba, Canada, n=982, and Egg Island, Alaska, USA, n=84) and one overwintering population (Cumberland Island, Georgia, USA, n=62). For Alaska and Georgia, time-invariant models were best-supported, giving annual survival estimates of 0.67 (95%C.I.: 0.58- 0.76) and 0.59 (95%C.I.: 0.49-0.67) respectively. Data from Manitoba supported a timedependent model: survival estimates varied from 1.00 to 0.36, with lowest estimates from recent years, supporting observations of local population decline. Seasonal survival analysis of the Georgia population indicated lower mortality during winter (monthly Φoverwinter: 0.959, 95%CI: 0.871-0.988; for 6 month period Φoverwinter: 0.780 (0.440-0.929)) than during combined breeding and migratory periods (monthly ΦBreeding+Migration: 0.879 (0.825-0.918); for 8 month ΦBreeding+Migration: 0356 (0.215-0.504)). I recommend, based on high resight rates, continued monitoring of survival of wintering populations, to determine potential range-wide population declines. Keywords: survival, longevity, mortality, shorebird, overwinter, breeding, migration, life cycle Author Keywords: life cycle, longevity, mortality, non-breeding, shorebird, survival
Tests of the Invasional Meltdown Hypothesis in invasive herbaceous plant species in southern Ontario
According to the Invasional Meltdown Hypothesis (IMH), invasive species may interact in their introduced range and facilitate future invasions. This study investigated the possibility that Alliaria petiolata, an invasive allelopathic herbaceous plant in Ontario, is facilitating invasions by additional alien species. Two allelopathic focal species were chosen for this study: the native Solidago canadensis and the invasive A. petiolata. Field surveys in southern Ontario that quantified plant biodiversity in plots that included one or both focal species revealed no support for the IMH, although fewer species co-existed with A. petiolata than with S. canadensis. A year-long recruitment experiment in Peterborough, Ontario, also produced results inconsistent with the IMH, although did provide some evidence that A. petiolata limited recruitment of other species. These results collectively show negative impacts on regional biodiversity by A. petiolata, even in the absence of an invasional meltdown. Author Keywords: allelopathy, Alliaria petiolata, co-occurrence surveys, invasional meltdown hypothesis, invasive species, Solidago canadensis
Habitat Characteristics, Density Patterns and Environmental Niches of Indo-Pacific Humpback Dolphins (Sousa chinensis) of the Pearl River Estuary and Eastern Taiwan Strait
The purpose of this thesis is to quantify the habitat characteristics, density patterns and environmental niches of two groups of Indo-Pacific humpback dolphins: Chinese white dolphins (CWD) of the Pearl River estuary (PRE), and Taiwanese white dolphins (TWD, =Taiwanese humpback dolphin, THD) found in the eastern Taiwan Strait (ETS). Much work has already been done on the habitat use of CWDs in parts of the PRE, so the purpose of my first two chapters was to advance knowledge of the TWD to a comparable level. Chapter 2 contains the first published description of the relatively shallow, inshore, estuarine habitat of the TWD. General environmental characteristics and observed group sizes were consistent with other populations of humpback dolphins, and group sizes were not correlated with the environmental variables measured during surveys. Chapter 3 investigated density patterns of TWDs, finding spatiotemporal heterogeneity across the study area. Humpback dolphin densities fluctuated from year to year, but some parts of the study area were consistently used more than others. Environmental characteristics again did not influence dolphin densities, though more dolphins than expected were sighted in waters adjacent to major land reclamations, which may be related to the location of these areas close to major rivers. In Chapter 4, niches of the TWD and CWDs found in the PRE were compared using species distribution models, which indicated significant niche overlap. This may be due to niche conservatism maintaining similar fundamental niches between the two groups since their historical split >10,000 years ago, or a result of the intrinsic biotic factors that influence occurrence data affecting the hypervolume dimensions of each realized niche in similar ways. Geographic predictions indicate that most of the TWD's range has likely been surveyed, and that there may be connectivity between PRE humpback dolphins and at least one neighbouring putative population due to continuous predicted suitable habitat in waters that remain poorly surveyed. Overall, my thesis demonstrates that density patterns may vary over time, but on a broad temporal scale, these two allopatric groups of Indo-Pacific humpback dolphins have similar habitat requirements in geographically isolated, but environmentally similar locations. Author Keywords: density, habitat, Indo-Pacific humpback dolphin, niche overlap, Sousa chinensis, species distribution model
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