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Microorganism

History

Evolution

Further information: Timeline of evolution

Single-celled microorganisms were the first forms of life to develop on Earth, approximately 34 billion years ago. Further evolution was slow, and for about 3 billion years in the Precambrian eon, all organisms were microscopic. So, for most of the history of life on Earth the only forms of life were microorganisms. Bacteria, algae and fungi have been identified in amber that is 220 million years old, which shows that the morphology of microorganisms has changed little since the triassic period.

Most microorganisms can reproduce rapidly and microbes such as bacteria can also freely exchange genes by conjugation, transformation and transduction between widely-divergent species. This horizontal gene transfer, coupled with a high mutation rate and many other means of genetic variation, allows microorganisms to swiftly evolve (via natural selection) to survive in new environments and respond to environmental stresses. This rapid evolution is important in medicine, as it has led to the recent development of ‘super-bugs’ pathogenic bacteria that are resistant to modern antibiotics.

Pre-Microbiology

The possibility that microorganisms exist was discussed for many centuries before their actual discovery in the 17th century. The earliest known idea to indicate the possibility of diseases spreading by yet unseen organisms was that of the Roman scholar Marcus Terentius Varro in a 1st century BC book titled On Agriculture in which he warns against locating a homestead near swamps:

and because there are bred certain minute creatures which cannot be seen by the eyes, which float in the air and enter the body through the mouth and nose and there cause serious diseases.

In The Canon of Medicine (1020), Ab Al ibn Sn (Avicenna) stated that bodily secretion is contaminated by foul foreign earthly bodies before being infected. He also hypothesized that tuberculosis and other diseases might be contagious, i.e. that they were infectious diseases, and used quarantine to limit their spread.

When the Black Death bubonic plague reached Andalusia in Spain, in the 14th century, Ibn Khatima wrote that infectious diseases were caused by contagious “minute bodies” that enter the human body. Later, in 1546, Girolamo Fracastoro proposed that epidemic diseases were caused by transferable seedlike entities that could transmit infection by direct or indirect contact, or even without contact over long distances.

All these early claims about the existence of microorganisms were speculative and were not based on any data or science. Microorganisms were neither proven, observed, nor correctly and accurately described until the 17th century. The reason for this was that all these early studies lacked the microscope.

History of microorganisms’ discovery

See also: History of biology

Antonie van Leeuwenhoek, the first microbiologist and the first to observe microorganisms using a microscope.

Anton van Leeuwenhoek was the one of the first people to observe microorganisms, using a microscope of his own design, and made one of the most important contributions to biology. Robert Hooke was the first to use a microscope to observe living things; his 1665 book Micrographia contained descriptions of plant cells.

Before Leeuwenhoek’s discovery of microorganisms in 1675, it had been a mystery why grapes could be turned into wine, milk into cheese, or why food would spoil. Leeuwenhoek did not make the connection between these processes and microorganisms, but using a microscope, he did establish that there were forms of life that were not visible to the naked eye. Leeuwenhoek’s discovery, along with subsequent observations by Lazzaro Spallanzani and Louis Pasteur, ended the long-held belief that life spontaneously appeared from non-living substances during the process of spoilage.

Lazzaro Spallanzani found that boiling broth would sterilise it and kill any microorganisms in it. He also found that new microorganisms could only settle in a broth if the broth was exposed to the air. Louis Pasteur expanded upon Spallanzani’s findings by exposing boiled broths to the air, in vessels that contained a filter to prevent all particles from passing through to the growth medium, and also in vessels with no filter at all, with air being admitted via a curved tube that would not allow dust particles to come in contact with the broth. By boiling the broth beforehand, Pasteur ensured that no microorganisms survived within the broths at the beginning of his experiment. Nothing grew in the broths in the course of Pasteur’s experiment. This meant that the living organisms that grew in such broths came from outside, as spores on dust, rather than spontaneously generated within the broth. Thus, Pasteur dealt the death blow to the theory of spontaneous generation and supported germ theory.

In 1767, Dr. J. Z. Holwell reported that Indian physicians at the time knew that microbes caused diseases: “They lay it down as a principle, that the immediate cause of the smallpox exists in the mortal part of every human and animal form; that the mediate (or second) acting cause, which stirs up the first, and throws it into a state of fermentation, is multitudes of imperceptible animalculae floating in the atmosphere; that these are the cause of all epidemical diseases, but more particularly of the small pox.”

In 1876, Robert Koch established that microbes can cause disease. He found that the blood of cattle who were infected with anthrax always had large numbers of Bacillus anthracis. Koch found that he could transmit anthrax from one animal to another by taking a small sample of blood from the infected animal and injecting it into a healthy one, and this caused the healthy animal to become sick. He also found that he could grow the bacteria in a nutrient broth, then inject it into a healthy animal, and cause illness. Based on these experiments, he devised criteria for establishing a causal link between a microbe and a disease and these are now known as Koch’s postulates. Although these postulates cannot be applied in all cases, they do retain historical importance to the development of scientific thought and are still being used today.

Classification and structure

Evolutionary tree showing the common ancestry of all three domains of life. Bacteria are colored blue, eukaryotes red, and archaea green. Relative positions of some phyla are shown around the tree.

Microorganisms can be found almost anywhere in the taxonomic organization of life on the planet. Bacteria and archaea are almost always microscopic, while a number of eukaryotes are also microscopic, including most protists, some fungi, as well as some animals and plants. Viruses are generally regarded as not living and therefore are not microbes, although the field of microbiology also encompasses the study of viruses.

Prokaryotes

Main article: Prokaryote

Prokaryotes are organisms that lack a cell nucleus and the other membrane bound organelles. They are almost always unicellular, although some species such as myxobacteria can aggregate into complex structures as part of their life cycle.

Consisting of two domains, bacteria and archaea, the prokaryotes are the most diverse and abundant group of organisms on Earth and inhabit practically all environments where some liquid water is available and the temperature is below +140 C. They are found in sea water, soil, air, animals’ gastrointestinal tracts, hot springs and even deep beneath the Earth’s crust in rocks. Practically all surfaces which have not been specially sterilized are covered by prokaryotes. The number of prokaryotes on Earth is estimated to be around five million trillion trillion, or 5 1030, accounting for at least half the biomass on Earth.

Bacteria

Main article: Bacteria

Staphylococcus aureus bacteria magnified about 10,000x

Bacteria are practically all invisible to the naked eye, with a few extremely rare exceptions, such as Thiomargarita namibiensis. They lack membrane-bound organelles, and can function and reproduce as individual cells, but often aggregate in multicellular colonies. Their genome is usually a single loop of DNA, although they can also harbor small pieces of DNA called plasmids. These plasmids can be transferred between cells through bacterial conjugation. Bacteria are surrounded by a cell wall, which provides strength and rigidity to their cells. They reproduce by binary fission or sometimes by budding, but do not undergo sexual reproduction. Some species form extraordinarily resilient spores, but for bacteria this is a mechanism for survival, not reproduction. Under optimal conditions bacteria can grow extremely rapidly and can double as quickly as every 10 minutes.

Archaea

Main article: Archaea

Archaea are also single-celled organisms that lack nuclei. In the past, the differences between bacteria and archaea were not recognised and archaea were classified with bacteria as part of the kingdom Monera. However, in 1990 the microbiologist Carl Woese proposed the three-domain system that divided living things into bacteria, archaea and eukaryotes. Archaea differ from bacteria in both their genetics and biochemistry. For example, while bacterial cell membranes are made from phosphoglycerides with ester bonds, archaean membranes are made of ether lipids.

Archaea were originally described in extreme environments, such as hot springs, but have since been found in all types of habitats. Only now are scientists beginning to realize how common archaea are in the environment, with crenarchaeota being the most common form of life in the ocean, dominating ecosystems below 150 m in depth. These organisms are also common in soil and play a vital role in ammonia oxidation.

Eukaryotes

Ostreococcus is the smallest known free living eukaryote with an average size of 0.8 m

Main article: Eukaryote

Most living things which are visible to the naked eye in their adult form are eukaryotes, including humans. However, a large number of eukaryotes are also microorganisms. Unlike bacteria and archaea, eukaryotes contain organelles such as the cell nucleus, the Golgi apparatus and mitochondria in their cells. The nucleus is an organelle which houses the DNA that makes up a cell’s genome. DNA itself is arranged in complex chromosomes. Mitochondria are organelles vital in metabolism as they are the site of the citric acid cycle and oxidative phosphorylation. They evolved from symbiotic bacteria and retain a remnant genome. Like bacteria, plant cells have cell walls, and contain organelles such as chloroplasts in addition to the organelles in other eukaryotes. Chloroplasts produce energy from light by photosynthesis, and were also originally symbiotic bacteria.

Unicellular eukaryotes are those eukaryotic organisms that consist of a single cell throughout their life cycle. This qualification is significant since most multicellular eukaryotes consist of a single cell called a zygote at the beginning of their life cycles. Microbial eukaryotes can be either haploid or diploid, and some organisms have multiple cell nuclei (see coenocyte). However, not all microorganisms are unicellular as some microscopic eukaryotes are made from multiple cells.

Protists

Main article: Protista

Of eukaryotic groups, the protists are most commonly unicellular and microscopic. This is a highly diverse group of organisms that are not easy to classify. Several algae species are multicellular protists, and slime molds have unique life cycles that involve switching between unicellular, colonial, and multicellular forms. The number of species of protozoa is uncertain, since we may have identified only a small proportion of the diversity in this group of organisms.

A microscopic mite Lorryia formosa.

Animals

Main article: Micro-animals

Mostly animals are multicellular, but some are too small to be seen by the naked eye. Microscopic arthropods include dust mites and spider mites. Microscopic crustaceans include copepods and the cladocera, while many nematodes are too small to be seen with the naked eye. Another particularly common group of microscopic animals are the rotifers, which are filter feeders that are usually found in fresh water. Micro-animals reproduce both sexually and asexually and may reach new habitats as eggs that survive harsh environments that would kill the adult animal. However, some simple animals, such as rotifers and nematodes, can dry out completely and remain dormant for long periods of time.

Fungi

Main article: Fungus

The fungi have several unicellular species, such as baker’s yeast (Saccharomyces cerevisiae) and fission yeast (Schizosaccharomyces pombe). Some fungi, such as the pathogenic yeast Candida albicans, can undergo phenotypic switching and grow as single cells in some environments, and filamentous hyphae in others. Fungi reproduce both asexually, by budding or binary fission, as well by producing spores, which are called conidia when produced asexually, or basidiospores when produced sexually.

Plants

Main article: Plant

The green algae are a large group of photosynthetic eukaryotes that include many microscopic organisms. Although some green algae are classified as protists, others such as charophyta are classified with embryophyte plants, which are the most familiar group of land plants. Algae can grow as single cells, or in long chains of cells. The green algae include unicellular and colonial flagellates, usually but not always with two flagella per cell, as well as various colonial, coccoid, and filamentous forms. In the Charales, which are the algae most closely related to higher plants, cells differentiate into several distinct tissues within the organism. There are about 6000 species of green algae.

Habitats and ecology

Microorganisms are found in almost every habitat present in nature. Even in hostile environments such as the poles, deserts, geysers, rocks, and the deep sea. Some types of microorganisms have adapted to the extreme conditions and sustained colonies; these organisms are known as extremophiles. Extremophiles have been isolated from rocks as much as 7 kilometres below the Earth’s surface, and it has been suggested that the amount of living organisms below the Earth’s surface may be comparable with the amount of life on or above the surface. Extremophiles have been known to survive for a prolonged time in a vacuum, and can be highly resistant to radiation, which may even allow them to survive in space. Many types of microorganisms have intimate symbiotic relationships with other larger organisms; some of which are mutually beneficial (mutualism), while others can be damaging to the host organism (parasitism). If microorganisms can cause disease in a host they are known as pathogens.

Extremophiles

Main article: Extremophile

Extremophiles are microorganisms which have adapted so that they can survive and even thrive in conditions that are normally fatal to most life-forms. For example, some species have been found in the following extreme environments:

Temperature: as high as 130 C (266 F), as low as 17 C (1.4 F)

Acidity/alkalinity: less than pH 0, up to pH 11.5

Salinity: up to saturation

Pressure: up to 1,000-2,000 atm, down to 0 atm (e.g. vacuum of space)

Radiation: up to 5kGy

Extremophiles are significant in different ways. They extend terrestrial life into much of the Earth’s hydrosphere, crust and atmosphere, their specific evolutionary adaptation mechanisms to their extreme environment can be exploited in bio-technology, and their very existence under such extreme conditions increases the potential for extraterrestrial life.

Soil microbes

The nitrogen cycle in soils depends on the fixation of atmospheric nitrogen. One way this can occur is in the nodules in the roots of legumes that contain symbiotic bacteria of the genera Rhizobium, Mesorhizobium, Sinorhizobium, Bradyrhizobium, and Azorhizobium.

Symbiotic microbes

Symbiotic microbes such as fungi and algae form an association in lichen. Certain fungi form mycorhizzal symbioses with trees that increase the supply of nutrients to the tree.

Importance

Microorganisms are vital to humans and the environment, as they participate in the Earth’s element cycles such as the carbon cycle and nitrogen cycle, as well as fulfilling other vital roles in virtually all ecosystems, such as recycling other organisms’ dead remains and waste products through decomposition. Microbes also have an important place in most higher-order multicellular organisms as symbionts. Many blame the failure of Biosphere 2 on an improper balance of microbes.

Use in food

Main article: Fermentation (food)

Microorganisms are used in brewing, winemaking, baking, pickling and other food-making processes.

They are also used to control the fermentation process in the production of cultured dairy products such as yogurt and cheese. The cultures also provide flavour and aroma, and inhibit undesirable organisms.

Use in water treatment

Main article: Sewage treatment

Specially-cultured microbes are used in the biological treatment of sewage and industrial waste effluent, a process known as bioaugmentation.

Use in energy

Main articles: Algae fuel, Cellulosic ethanol, and Ethanol fermentation

Microbes are used in fermentation to produce ethanol, and in biogas reactors to produce methane. Scientists are researching the use of algae to produce liquid fuels, and bacteria to convert various forms of agricultural and urban waste into usable fuels.

Use in science

Microbes are also essential tools in biotechnology, biochemistry, genetics, and molecular biology. The yeasts (Saccharomyces cerevisiae) and fission yeast (Schizosaccharomyces pombe) are important model organisms in science, since they are simple eukaryotes that can be grown rapidly in large numbers and are easily manipulated. They are particularly valuable in genetics, genomics and proteomics. Microbes can be harnessed for uses such as creating steroids and treating skin diseases. Scientists are also considering using microbes for living fuel cells, and as a solution for pollution.

Use in warfare

Main article: Biological warfare

In the Middle Ages, diseased corpses were thrown into castles during sieges using catapults or other siege engines. Individuals near the corpses were exposed to the deadly pathogen and were likely to spread that pathogen to others.

Importance in human health

Human digestion

Further information: Human flora#Human bacterial flora and human health

Microorganisms can form an endosymbiotic relationship with other, larger organisms. For example, the bacteria that live within the human digestive system contribute to gut immunity, synthesise vitamins such as folic acid and biotin, and ferment complex indigestible carbohydrates.

Diseases and immunology

Main article: Pathogenic microbes

Microorganisms are the cause of many infectious diseases. The organisms involved include pathogenic bacteria, causing diseases such as plague, tuberculosis and anthrax; protozoa, causing diseases such as malaria, sleeping sickness and toxoplasmosis; and also fungi causing diseases such as ringworm, candidiasis or histoplasmosis. However, other diseases such as influenza, yellow fever or AIDS are caused by pathogenic viruses, which are not usually classified as living organisms and are not therefore microorganisms by the strict definition. As of 2007, no clear examples of archaean pathogens are known, although a relationship has been proposed between the presence of some methanogens and human periodontal disease.

Importance in ecology

Further information: Decomposition

Microbes are critical to the processes of decomposition required to cycle nitrogen and other elements back to the natural world.

Hygiene

Main article: Hygiene

Hygiene is the avoidance of infection or food spoiling by eliminating microorganisms from the surroundings. As microorganisms, particularly bacteria, are found practically everywhere, this means in most cases the reduction of harmful microorganisms to acceptable levels. However, in some cases it is required that an object or substance be completely sterile, i.e. devoid of all living entities and viruses. A good example of this is a hypodermic needle.

In food preparation microorganisms are reduced by preservation methods (such as the addition of vinegar), clean utensils used in preparation, short storage periods or by cool temperatures. If complete sterility is needed, the two most common methods are irradiation and the use of an autoclave, which resembles a pressure cooker.

There are several methods for investigating the level of hygiene in a sample of food, drinking water, equipment etc. Water samples can be filtrated through an extremely fine filter. This filter is then placed in a nutrient medium. Microorganisms on the filter then grow to form a visible colony. Harmful microorganisms can be detected in food by placing a sample in a nutrient broth designed to enrich the organisms in question. Various methods, such as selective media or PCR, can then be used for detection. The hygiene of hard surfaces, such as cooking pots, can be tested by touching them with a solid piece of nutrient medium and then allowing the microorganisms to grow on it.

There are no conditions where all microorganisms would grow, and therefore often several different methods are needed. For example, a food sample might be analyzed on three different nutrient mediums designed to indicate the presence of “total” bacteria (conditions where many, but not all, bacteria grow), molds (conditions where the growth of bacteria is prevented by e.g. antibiotics) and coliform bacteria (these indicate a sewage contamination).

See also

Biological warfare

Biology

Culture collection

Microbial intelligence

Nanobacterium

Petri dish

Prokaryote

Soil contamination

Staining

Virus

Bacterium

Protozoa

Fungi

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^ Some Psychrophilic bacteria can grow at 17 C (1.4 F), and can survive near absolute zero.

^ Picrophilus can grow at pH -0.06.

^ The alkaliphilic bacteria Bacillus alcalophilus can grow at up to pH 11.5.

^ Dyall-Smith, Mike, HALOARCHAEA, University of Melbourne. See also Haloarchaea.

^ The piezophilic bacteria Halomonas salaria requires a pressure of 1,000 atm; nanobes, a speculative organism, have been reportedly found in the earth’s crust at 2,000 atm.

^ See Deinococcus radiodurans

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^ Gray, N.F. (2004). Biology of Wastewater Treatement. Imperial College Press. p. 1164. ISBN 1-860-94332-2. 

^ Kitani, Osumu and Carl W. Hall (1989). Biomass Handbook. Taylor & Francis US. p. 256. ISBN 2-881-24269-3. 

^ Pimental, David (2007). Food, Energy, and Society. CRC Press. p. 289. ISBN 1-420-04667-5. 

^ Tickell, Joshua et al. (2000). From the Fryer to the Fuel Tank: The Complete Guide to Using Vegetable Oil as an Alternative Fuel. Biodiesel America. p. 53. ISBN 0-970-72270-2. 

^ Inslee, Jay et al. (2008). Apollo’s Fire: Igniting America’s Clean Energy Economy. Island Press. p. 157. ISBN 1-597-26175-0. 

^ Castrillo JI, Oliver SG (2004). “Yeast as a touchstone in post-genomic research: strategies for integrative analysis in functional genomics”. J. Biochem. Mol. Biol. 37 (1): 93106. PMID 14761307. http://www.jbmb.or.kr/fulltext/jbmb/view.php?vol=37&page=93. 

^ Suter B, Auerbach D, Stagljar I (2006). “Yeast-based functional genomics and proteomics technologies: the first 15 years and beyond”. BioTechniques 40 (5): 62544. doi:10.2144/000112151. PMID 16708762. 

^ Sunnerhagen P (2002). “Prospects for functional genomics in Schizosaccharomyces pombe”. Curr. Genet. 42 (2): 7384. doi:10.1007/s00294-002-0335-6. PMID 12478386. 

^ Soni, S.K. (2007). Microbes: A Source of Energy for 21st Century. New India Publishing. ISBN 8-189-42214-6. 

^ Moses, Vivian et al. (1999). Biotechnology: The Science and the Business. CRC Press. p. 563. ISBN 9-057-02407-1. 

^ Langford, Roland E. (2004). Introduction to Weapons of Mass Destruction: Radiological, Chemical, and Biological. Wiley-IEEE. p. 140. ISBN 0-471-46560-7. 

^ O’Hara A, Shanahan F (2006). “The gut flora as a forgotten organ”. EMBO Rep 7 (7): 68893. doi:10.1038/sj.embor.7400731. PMID 16819463. 

^ Eckburg P, Lepp P, Relman D (2003). “Archaea and their potential role in human disease”. Infect Immun 71 (2): 5916. doi:10.1128/IAI.71.2.591-596.2003. PMID 12540534. 

^ Lepp P, Brinig M, Ouverney C, Palm K, Armitage G, Relman D (2004). “Methanogenic Archaea and human periodontal disease”. Proc Natl Acad Sci USA 101 (16): 617681. doi:10.1073/pnas.0308766101. PMID 15067114. 

External links

Our Microbial Planet A free poster from the National Academy of Sciences about the positive roles of microbes.

“Uncharted Microbial World: Microbes and Their Activities in the Environment” Report from the American Academy of Microbiology

Understanding Our Microbial Planet: The New Science of Metagenomics A 20-page educational booklet providing a basic overview of metagenomics and our microbial planet.

Tree of Life Eukaryotes

Microbe News from Genome News Network

Microbes Patent List Microbes Related Patents

Medical Microbiology On-line textbook

Through the microscope: A look at all things small On-line microbiology textbook by Timothy Paustian and Gary Roberts, University of Wisconsin-Madison

MicrobeID.com Online Bacteria Identification Key and Probabilistic Identification Databases

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Sonic Goes Spore Adventure 7 Beta Test

Spore Spore Journal by CafePress $11 Scribble important stuff – lyrics, recipes, addresses, and more. Our Wire-O bound, 160 page journal has your choice of papers and measures 5 x 8, a handy on-the-go size to fit in your backpack. Get creative and let the muse flow. Back cove Spore Journal Scribble important stuff – lyrics, recipes, addresses, and more. Our Wire-O bound, 160 page journal has your choice of papers and measures 5 x 8, a handy on-the-go size to fit in your backpack. Get creative and let the muse flow. Back cove

Milky Spore 10 Ounce $22.99 MILKY SPORE GRUB & BEETLE CONTROL There is no better way to eliminate these unwanted pests! Produced to USDA standards, MILKY SPORE is the safest material ever developed for control over the larvae (or Grubs) of Japanese Beetles. Grubs do damage to your lawn and attract digging vagrants such as moles and voles. Milky Spore is a naturally occurring host specific bacterium (Bacillus popillae-Dutky). This product is lethal to a familiar destructive summer-time pest. It targets and discriminately works to attack the white grubs of Japanese Beetles. Milky Spore Frequently Asked Questions What is Milky Spore? Milky Spore is a naturally occurring bacterium that affects Japanese Beetle, June Beetle and certain other common lawn grubs. How fast does the Spore work? Milky Spore begins working upon application wherever grubs are feeding. Warm climates can achieve complete control in 6 months to 1 year. Colder areas may require 1 to 2 years. How long does it last? Once established in the lawn, Milky Spore lasts 15 to 20 years! Is it safe to use Around Children & Pets? Yes, Milky Spore is not a poison. It will not affect humans, pets, birds, fish or other beneficial insects, ponds, streams or well water. Will heavy rain, freezing rain, or snow affecy the Spore? No. Once applied Milky Spore is not affected by environmental conditions. Will fertilizer affect the Spore? No. Milky Spore is not affected by lawn chemicals. They may all be applied at the same time. Can I use Milky Spore in the vegetable garden? Absolutely! Milky Spore is harmless to food crops. Milky Spore may also be used on pasture land. Can I mix the Spore in water and spray it? No. Milky Spore must be applied directly in its original dry state. Can I cut my grass after applying the Spore? No. Do not mow your grass until the Spore has been watered into the soil by rain or hose for 15 minutes or longer. What if I treat my yard and my neighbor doesn’t? Your lawn will be protected even if surrounded by untreated property. Spore will eventually spread by natural movement. The Milky Spore Process! The adult beetle feeds on fruit, flower, shrubs, garden plants and the foliage of some field crops. At the immature beetle stage, the grub enjoys feeding on the roots of grass and other vegetation to include stems of plants. Turf inoculation treatments / applications with MILKY SPORE puts in place an on-guard protective blanket on your lawn. Considered the weakest link in the chain and the most vulnerable point to introduce an infection, resident spores in treated turf are swallowed by grubs during their normal pattern of feeding; this starts the demise of healthy grubs. Milky Spore disease then begins to cripple the grub, and within the next 7-21 days will eventually die. As the grub decomposes, it releases billions of new spores. Available in 2 Sizes 10 Ounce and 2.5 Pounds Milky Spore Japanese Beetle Grub Control Application Directions Apply one level teaspoon of Milky Spore powder on top of grass every 3

Milky Spore 40 Ounce $68.99 MILKY SPORE GRUB & BEETLE CONTROL There is no better way to eliminate these unwanted pests! Produced to USDA standards, MILKY SPORE is the safest material ever developed for control over the larvae (or Grubs) of Japanese Beetles. Grubs do damage to your lawn and attract digging vagrants such as moles and voles. Milky Spore is a naturally occurring host specific bacterium (Bacillus popillae-Dutky). This product is lethal to a familiar destructive summer-time pest. It targets and discriminately works to attack the white grubs of Japanese Beetles. Milky Spore Frequently Asked Questions What is Milky Spore? Milky Spore is a naturally occurring bacterium that affects Japanese Beetle, June Beetle and certain other common lawn grubs. How fast does the Spore work? Milky Spore begins working upon application wherever grubs are feeding. Warm climates can achieve complete control in 6 months to 1 year. Colder areas may require 1 to 2 years. How long does it last? Once established in the lawn, Milky Spore lasts 15 to 20 years! Is it safe to use Around Children & Pets? Yes, Milky Spore is not a poison. It will not affect humans, pets, birds, fish or other beneficial insects, ponds, streams or well water. Will heavy rain, freezing rain, or snow affecy the Spore? No. Once applied Milky Spore is not affected by environmental conditions. Will fertilizer affect the Spore? No. Milky Spore is not affected by lawn chemicals. They may all be applied at the same time. Can I use Milky Spore in the vegetable garden? Absolutely! Milky Spore is harmless to food crops. Milky Spore may also be used on pasture land. Can I mix the Spore in water and spray it? No. Milky Spore must be applied directly in its original dry state. Can I cut my grass after applying the Spore? No. Do not mow your grass until the Spore has been watered into the soil by rain or hose for 15 minutes or longer. What if I treat my yard and my neighbor doesn’t? Your lawn will be protected even if surrounded by untreated property. Spore will eventually spread by natural movement. The Milky Spore Process! The adult beetle feeds on fruit, flower, shrubs, garden plants and the foliage of some field crops. At the immature beetle stage, the grub enjoys feeding on the roots of grass and other vegetation to include stems of plants. Turf inoculation treatments / applications with MILKY SPORE puts in place an on-guard protective blanket on your lawn. Considered the weakest link in the chain and the most vulnerable point to introduce an infection, resident spores in treated turf are swallowed by grubs during their normal pattern of feeding; this starts the demise of healthy grubs. Milky Spore disease then begins to cripple the grub, and within the next 7-21 days will eventually die. As the grub decomposes, it releases billions of new spores. Available in 2 Sizes 10 Ounce and 2.5 Pounds Milky Spore Japanese Beetle Grub Control Application Directions Apply one level teaspoon of Milky Spore powder on top of grass every 3

Milky Spore 50 LB Drum $1249.99 MILKY SPORE GRUB & BEETLE CONTROL There is no better way to eliminate these unwanted pests! Produced to USDA standards, MILKY SPORE is the safest material ever developed for control over the larvae (or Grubs) of Japanese Beetles. Grubs do damage to your lawn and attract digging vagrants such as moles and voles. Milky Spore is a naturally occurring host specific bacterium (Bacillus popillae-Dutky). This product is lethal to a familiar destructive summer-time pest. It targets and discriminately works to attack the white grubs of Japanese Beetles. Milky Spore Frequently Asked Questions What is Milky Spore? Milky Spore is a naturally occurring bacterium that affects Japanese Beetle, June Beetle and certain other common lawn grubs. How fast does the Spore work? Milky Spore begins working upon application wherever grubs are feeding. Warm climates can achieve complete control in 6 months to 1 year. Colder areas may require 1 to 2 years. How long does it last? Once established in the lawn, Milky Spore lasts 15 to 20 years! Is it safe to use Around Children & Pets? Yes, Milky Spore is not a poison. It will not affect humans, pets, birds, fish or other beneficial insects, ponds, streams or well water. Will heavy rain, freezing rain, or snow affecy the Spore? No. Once applied Milky Spore is not affected by environmental conditions. Will fertilizer affect the Spore? No. Milky Spore is not affected by lawn chemicals. They may all be applied at the same time. Can I use Milky Spore in the vegetable garden? Absolutely! Milky Spore is harmless to food crops. Milky Spore may also be used on pasture land. Can I mix the Spore in water and spray it? No. Milky Spore must be applied directly in its original dry state. Can I cut my grass after applying the Spore? No. Do not mow your grass until the Spore has been watered into the soil by rain or hose for 15 minutes or longer. What if I treat my yard and my neighbor doesn’t? Your lawn will be protected even if surrounded by untreated property. Spore will eventually spread by natural movement. The Milky Spore Process! The adult beetle feeds on fruit, flower, shrubs, garden plants and the foliage of some field crops. At the immature beetle stage, the grub enjoys feeding on the roots of grass and other vegetation to include stems of plants. Turf inoculation treatments / applications with MILKY SPORE puts in place an on-guard protective blanket on your lawn. Considered the weakest link in the chain and the most vulnerable point to introduce an infection, resident spores in treated turf are swallowed by grubs during their normal pattern of feeding; this starts the demise of healthy grubs. Milky Spore disease then begins to cripple the grub, and within the next 7-21 days will eventually die. As the grub decomposes, it releases billions of new spores. Available in 2 Sizes 10 Ounce and 2.5 Pounds Milky Spore Japanese Beetle Grub Control Application Directions Apply one level teaspoon of Milky Spore powder on top of grass every 3

Milky Spore Grub Control Dispenser Tube $6.19 MILKY SPORE GRUB & BEETLE CONTROL One Application Lasts 15 years! There is no better way to eliminate these unwanted pests! Produced to USDA standards, MILKY SPORE is the safest material ever developed for control over the larvae (or Grubs) of Japanese Beetles. Grubs do damage to your lawn and attract digging vagrants such as moles and voles. Milky Spore is a naturally occurring host specific bacterium (Bacillus popillae-Dutky). This product is lethal to a familiar destructive summer-time pest. It targets and discriminately works to attack the white grubs of Japanese Beetles. Milky Spore is not harmful to beneficial insects, birds, bees, pets or man. The product is approved and registered with EPA, Milky Spore will not affect wells, ponds or streams. Interested in learning how to identify a white grub infestation? Milky Spore Japanese Beetle Grub Control Application Directions : Apply one level teaspoon of Milky Spore powder on top of grass every 3-4 feet. Application should be in a grid pattern every 3-4 Feet. Apply by hand or purchase our applicator shown to the right Water lightly after application to soak spores into soil Areas between spots will be inoculated naturally How Much Do I Need: Use 10 ounces per 2500 square feet (50 ft. by 50 ft.) Use 40 ounces per 10000 square feet (100 ft. by 100 ft.) Milky Spore Frequently Asked Questions: What is Milky Spore? Milky Spore is a naturally occurring bacterium that affects Japanese Beetle, June Beetle and certain other common lawn grubs. How fast does the Spore work? Milky Spore begins working upon application wherever grubs are feeding. Warm climates can achieve complete control in 6 months to 1 year. Colder areas may require 1 to 2 years. How long does it last? Once established in the lawn, Milky Spore lasts 15 to 20 years! Is it safe to use Around Children & Pets? Yes, Milky Spore is not a poison. It will not affect humans, pets, birds, fish or other beneficial insects, ponds, streams or well water. Will heavy rain, freezing rain, or snow affecy the Spore? No. Once applied Milky Spore is not affected by environmental conditions. Will fertilizer affect the Spore? No. Milky Spore is not affected by lawn chemicals. They may all be applied at the same time. Can I use Milky Spore in the vegetable garden? Absolutely! Milky Spore is harmless to food crops. Milky Spore may also be used on pasture land. Can I mix the Spore in water and spray it? No. Milky Spore must be applied directly in its original dry state. Can I cut my grass after applying the Spore? No. Do not mow your grass until the Spore has been watered into the soil by rain or hose for 15 minutes or longer. What if I treat my yard and my neighbor doesn’t? Your lawn will be protected even if surrounded by untreated property. Spore will eventually spread by natural movement. The Milky Spore Process! The adult beetle feeds on fruit, flower, shrubs, garden plants and the foliage of some field crops. At the immature beetle stage, the grub enjoys f

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Region Test Hotel 5 $391 Region Test Hotel 5 is a business friendly property located in Region Test. Property Features. Region Test Hotel 5 features 2 beach bars, 2 poolside bars, 3 bars/lounges, and a restaurant. A bar/lounge is open for drinks. Room service is available 24 hours a day. The property serves a complimentary continental breakfast in the restaurant on weekdays. Guests can enjoy a complimentary dinner reception in the early evening, available Monday, Wednesday, and Friday. Recreational amenities include 2 heated indoor swimming pools. Also located on site are a children’s pool, a fitness facility, and a children’s club. This 4.5 star property has a business center and offers business services. Wireless Internet access is available in public areas. This Region Test property has event space consisting of banquet facilities, conference/meeting rooms, and a ballroom. The property has a roundtrip airport shuttle, which is complimentary. Guest parking is complimentary. Additional property amenities include a marina, a library, and gift shops/newsstands. Guestrooms. 73 air conditioned guestrooms at Region Test Hotel 5 feature coffee/tea makers and safes. Furnishings include queen sofa beds and desks. Bathrooms feature shower/tub combinations, makeup/shaving mirrors, complimentary toiletries, and hair dryers. Wired high speed Internet access is complimentary. In addition to complimentary weekday newspapers, guestrooms offer direct dial phones as well as free local calls (restrictions may apply). Satellite television is provided. Also included are irons/ironing boards and clock radios. Official United States of America Classification provided by ANBBAANBBA assigns an official classification for accommodations in United States of America. For this type of property the ANBBA assigns ratings from Categoria I (highest classification) to Categoria IV SUP (lowest classification). This Rural House property is awarded the following classification: Catergoria I SUP. For the benefit of our customers, we have provided a rating based on our rating system. Notifications and Fees:Guests requiring a visa to enter Russia may be able to obtain help from the hotel where they will be staying. Guests should contact the hotel using the contact information on the booking confirmation. Hotels may charge for providing visa assistance even if guests subsequently cancel their reservation. All arrangements including any applicable charges are made directly between guests and the hotel. In addition, guests will also be liable for any applicable cancellation charges. Couples wishing to share a room must provide proof of marriage. Under Vietnamese law, a guest who is not a Vietnamese citizen cannot share a room with a Vietnamese citizen without proof of marriage to each other. Otherwise, a second room must be booked. Guests must transfer via boat or floatplane to the property. Guests must contact the property at least 72 hours before travel, using the contact information on the confirmation

EQC Test Hotel (PDP) $393.24 EQC Test Hotel (PDP) is located in Region Test. Property Features. EQC Test Hotel (PDP) features a coffee shop/caf? and a bar/lounge. The property serves Hot and cold buffet breakfasts (surcharges apply). Guests can enjoy a complimentary dinner reception in the late afternoon on select days. Recreational amenities include an outdoor pool, an indoor pool, and a children’s pool. The property’s full service health spa has body treatments and massage/treatment rooms. This property has a business center and offers small meeting rooms, limo/town car service, and audio visual equipment. Complimentary high speed (wired) Internet access is available in public areas. This Region Test property has event space consisting of a ballroom and exhibit space. For a surcharge, shuttle services include a roundtrip airport shuttle and an area shuttle. Additional property amenities include a concierge desk, an arcade/game room, and a nightclub. Guestrooms. Air conditioned guestrooms at EQC Test Hotel (PDP) feature minibars and CD players. Rooms are all accessible via exterior corridors. Beds come with premium bedding. All rooms include separate sitting areas and desks. Kitchens or kitchenettes include full sized refrigerators/freezers, stovetops, microwaves, and dishwashers. Guestrooms share bathrooms; amenities include double sinks, televisions, and bathrobes. Wired high speed Internet access is available for a surcharge. In addition to complimentary newspapers and laptop compatible safes, guestrooms offer cordless phones as well as free local calls (restrictions may apply). Televisions have cable channels. Also included are complimentary bottled water and ceiling fans. Guests may request irons/ironing boards, hair dryers, and extra towels/bedding. Official United States of America Rating provided by Enjoy EnglandEnjoy England assigns an official star rating for accommodations in United States of America. This Self Contained & Serviced property is rated 3 daisies. Notifications and Fees:Guests requiring a visa to enter Russia may be able to obtain help from the hotel where they will be staying. Guests should contact the hotel using the contact information on the booking confirmation. Hotels may charge for providing visa assistance even if guests subsequently cancel their reservation. All arrangements including any applicable charges are made directly between guests and the hotel. In addition, guests will also be liable for any applicable cancellation charges. Couples wishing to share a room must provide proof of marriage. Under Vietnamese law, a guest who is not a Vietnamese citizen cannot share a room with a Vietnamese citizen without proof of marriage to each other. Otherwise, a second room must be booked. To arrange required transfers to the property, guests must contact the property before travel using the contact information on the booking confirmation. A mandatory fee for lunch on 18 April is included in the total price displayed Minimum Spring Break check

EQC Test Hotel $198 EQC Test Hotel is located in Region Test. Property Features. Recreational amenities include a children’s pool. Those traveling on business have access to a business center at this property. The property serves a complimentary full breakfast in the pool bar on weekends. For a surcharge, the property offers a roundtrip airport shuttle 5:00. Additional property amenities include a casino, a concierge desk, and barbecue grills. Guestrooms. Air conditioned guestrooms at EQC Test Hotel feature fireplaces and minibars. Rooms are all accessible via exterior corridors. All accommodations have balconies. All rooms include separate sitting areas and desks. Kitchens include microwaves, refrigerators, dishwashers, and coffee/tea makers. Bathrooms feature showers with jetted bathtubs and handheld showerheads. They also offer televisions, bidets, and makeup/shaving mirrors. Wireless Internet access is available. In addition to complimentary weekday newspapers and safes, guestrooms offer cordless phones with voice mail, as well as free local calls (restrictions may apply). 50/70 cm high definition televisions have premium satellite channels, DVD players, and video game consoles. Rooms also include CD players and washers/dryers. Guests may request extra towels/bedding and wake up calls. Housekeeping is available daily. Official United States of America Classification provided by Hellenic Chamber of HotelsHellenic Chamber of Hotels assigns an official classification for accommodations in United States of America. For this type of property the Hellenic Chamber of Hotels assigns ratings from Deluxe Class (highest classification) to Standard Class (lowest classification). This Gite property is awarded the following classification: Boutique Class. For the benefit of our customers, we have provided a rating based on our rating system. Notifications and Fees:Guests requiring a visa to enter Russia may be able to obtain help from the hotel where they will be staying. Guests should contact the hotel using the contact information on the booking confirmation. Hotels may charge for providing visa assistance even if guests subsequently cancel their reservation. All arrangements including any applicable charges are made directly between guests and the hotel. In addition, guests will also be liable for any applicable cancellation charges. Clothing is optional at this property. Couples wishing to share a room must provide proof of marriage. Under Vietnamese law, a guest who is not a Vietnamese citizen cannot share a room with a Vietnamese citizen without proof of marriage to each other. Otherwise, a second room must be booked. To arrange required transfers to the property, guests must contact the property before travel using the contact information on the booking confirmation. Minimum Spring Break check in age is 18 years old. The Japanese Ministry of Health, Labour, and Welfare requires all international visitors to submit their passport number and nationality when registe

EQC Test Hotel (PPP) $375 EQC Test Hotel (PPP) is located in Region Test. Property Features. EQC Test Hotel (PPP) features a restaurant and a bar/lounge. Room service is available 24 hours a day. The villa property serves Continental breakfasts (surcharges apply). Recreational amenities include a golf course. Also located on site are an outdoor pool, an indoor pool and a steam room. Spa amenities include spa services and body treatments. Wireless and wired high speed Internet access is available in public areas (surcharges apply). In addition to a roundtrip airport shuttle, other complimentary shuttle services include an area shuttle and a train station pick up service. Tour/ticket assistance and translation services are available. Additional property amenities include a casino, a concierge desk, and barbecue grills. Guestrooms. Air conditioned guestrooms at EQC Test Hotel (PPP) feature minibars and CD players. All accommodations have balconies. Beds come with premium bedding. All rooms include separate sitting areas and desks. Refrigerators, microwaves, and coffee/tea makers are offered. Guestrooms share bathrooms; amenities include double sinks, televisions, and bidets. Wired high speed Internet access is available for a surcharge. In addition to safes, guestrooms offer direct dial phones as well as free local calls (restrictions may apply). 100 150 cm plasma televisions have DVD players. Also included are complimentary bottled water and bathrobes. Housekeeping is offered daily and guests may request hair dryers. Official United States of America Classification provided by United States of America Ministry of Tourism and CultureUnited States of America Ministry of Tourism and Culture assigns an official classification for accommodations in United States of America. This Guest & Hosted property is awarded the following classification: Boutique Class. Notifications and Fees:A mandatory fee for brunch is included in the total price displayedMinimum Spring Break check in age is 18 years old. Some properties have extra fees for amenities or services that may apply even if you do not use them. Government fees or taxes also may be charged to you when you check in or check out. This property told us they will charge you for the following: A fee for breakfast on 1 September: USD 15 20 per person A tax is imposed by the city: USD 25 30 per accommodation, per stayTransfer service fee per adult: USD 25/50Spa fee: USD 12.50 per stay We have included all charges provided to us by the property. However, charges can vary, for example, based on length of stay or the room you book.The following fees and deposits are charged by the property at time of service, check in, or check out. Early check in fee: USD 15/20Fee for high speed Internet (wired) in business center: USD 15 (for 1/4 hours, rates may vary)Fee for wireless Internet in business center: USD 18 (for 1 3 hours, rates may vary)Fee for high speed Internet (wired) in public areas: USD 20 (for 1/3 hours, rates may vary)Fee

ARI Test Hotel $225 ARI Test Hotel is a beachfront hotel located in Region Test’s midtown area. Hotel Features. Recreational amenities include 2 indoor swimming pools and 3 outdoor swimming pools. Also located on site are outdoor tennis courts, an indoor tennis court, and a children’s club. This hotel has a 24 hour business center and offers audio visual equipment. This Region Test property has event space consisting of a conference center, conference/meeting rooms, and a ballroom. The hotel serves a complimentary continental breakfast each morning in the caf?. In addition to a cruise ship terminal shuttle, other complimentary shuttle services include a ferry terminal shuttle, a shopping center shuttle, and a train station pick up service. Guest parking is limited, and available on a limited first come, first served basis (surcharge). Nearby parking is also available for a surcharge. Other property amenities at this Art Deco property include a concierge desk, barbecue grills, and an arcade/game room. Guestrooms. 200 air conditioned guestrooms at ARI Test Hotel feature fireplaces and private pools. Rooms are all accessible via exterior corridors. Patios offer golf course, beach, or ocean views. Beds come with Select Comfort mattresses, triple sheeting, down comforters, and down blankets. In addition to separate dining areas, these individually furnished and decorated rooms include desks and ergonomic chairs. Kitchens include full sized refrigerators/freezers, stovetops, microwaves, and dishwashers. Bathrooms feature separate bathtubs and showers with jetted bathtubs, handheld showerheads, and rainfall showerheads. They also offer double sinks, televisions, and bidets. Wired high speed and wireless Internet access is complimentary. In addition to complimentary weekday newspapers and laptop compatible safes, guestrooms offer cordless phones with voice mail, as well as complimentary local and international calls (restrictions may apply). 50 inch plasma televisions have premium cable channels, first run movies, and free movie channels. Also included are private plunge pools and iPod docking stations. Guests may request in room massages, hair dryers, and hypo allergenic bedding. Housekeeping is available weekly. Guestrooms are all non smoking. Notifications and Fees:Clothing is optional at this property. Local laws may restrict unmarried guests from sharing rooms. Guests are responsible for providing proof of marriage, if requested by the hotel. Couples wishing to share a room must provide proof of marriage. This property’s policy is to refuse certain bookings for the purpose of group events or parties, including pre wedding stag/bachelor and hen/bachelorette parties. Guests must transfer via airplane to the property. Guests must contact the property at least 24 hours before travel, using the contact information on the confirmation received after booking. Minimum Spring Break check in age is 25 years old. Children 3 years old and younger are not permitted from Febru

TEST HOTEL DO NOT RESERVE IT $211.27 Hotel Year Built 2011 NBR OF NON SMOKING ROOMS 20NBR OF ROOMS FOR PHYSICALLY CHALLENGED GUESTS 10A2T MAX OCCUPANCY 5 MAX NBR OF ROLLAWAYS 1 ROOM DESCRIPTION 1 KING SIZE BED WITH OCEAN VIEW ROOM AMENITIES 120 AC / 220 AC / AM/FM ALARM CLOCK / BATH TUB / CABLE/SATELLITE TV / CLOTHES PRESS / COFFEE/TEA MAKER / COLOR TV / CONNECTING ROOM / DIRECT DIAL TELEPHONE / EASY CHAIR / FREE LOCAL TELEPHONE CALLS / FREE NEWSPAPER / HAIRDRYER / HANDICAPPED ROOM/FACILITIES / IRON/IRONING BOARD / KITCHEN / MICROWAVE / MINI BAR / NO SMOKING ROOM / PHONE IN BATHROOM / REFRIGERATOR / REFRIGERATOR ICE MAKER / ROOM GAME SYSTEM / SAFE / TELEVISION / WORK DESK W/ LAMP TECHNOLOGY CORDLESS PHONE / DATAPORT / FAX MACHINE / HIGH SPEED INTERNET ACCESS / VOICE MAILA3T MAX OCCUPANCY 3 MAX NBR OF ROLLAWAYS 1 ROOM DESCRIPTION STANDARD ROOM WITH 3 SINGLE BEDS ROOM AMENITIES 120 AC / 220 AC / AM/FM ALARM CLOCK / BATH TUB / CABLE/SATELLITE TV / CLOTHES PRESS / COFFEE/TEA MAKER / COLOR TV / CONNECTING ROOM / DIRECT DIAL TELEPHONE / EASY CHAIR / FREE LOCAL TELEPHONE CALLS / FREE NEWSPAPER / HAIRDRYER / HANDICAPPED ROOM/FACILITIES / IRON/IRONING BOARD / KITCHEN / MICROWAVE / MINI BAR / NO SMOKING ROOM / PHONE IN BATHROOM / REFRIGERATOR / REFRIGERATOR ICE MAKER / ROOM GAME SYSTEM / SAFE / TELEVISION / WORK DESK W/ LAMP TECHNOLOGY CORDLESS PHONE / DATAPORT / FAX MACHINE / HIGH SPEED INTERNET ACCESS / VOICE MAILB1D MAX OCCUPANCY 5 MAX NBR OF ROLLAWAYS 1 ROOM DESCRIPTION 1 DB BED TEST POUR RATE MANAGER ROOM AMENITIES 120 AC / 220 AC / AM/FM ALARM CLOCK / BATH TUB / CABLE/SATELLITE TV / CLOTHES PRESS / COFFEE/TEA MAKER / COLOR TV / CONNECTING ROOM / DIRECT DIAL TELEPHONE / EASY CHAIR / FREE LOCAL TELEPHONE CALLS / FREE NEWSPAPER / HAIRDRYER / HANDICAPPED ROOM/FACILITIES / IRON/IRONING BOARD / KITCHEN / MICROWAVE / MINI BAR / NO SMOKING ROOM / PHONE IN BATHROOM / REFRIGERATOR / REFRIGERATOR ICE MAKER / ROOM GA

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Expedia Connect Test Hotel 1 $134.32 Located in Region Test, Expedia Connect Test Hotel 1 is a beachfront spa hotel in the historical district. Hotel Features. This Region Test hotel has a private beach. In addition to 10 restaurants, Expedia Connect Test Hotel 1 offers a coffee shop/caf? and a grocery/convenience store. A swim up bar, a poolside bar, and a bar/lounge are open for drinks. Room service is available during limited hours. A complimentary continental breakfast is served on weekdays. Guests can enjoy a complimentary manager reception in the late afternoon, available each Thursday. Recreational amenities include a waterpark with a lazy river and a waterslide, as well as 5 outdoor swimming pools. Also located on site are a golf course, an indoor pool, a health club, and outdoor tennis courts. The property’s full service health spa has massage/treatment rooms. This hotel has a 24 hour business center and offers small meeting rooms, secretarial services, and a technology helpdesk. Wireless and wired high speed Internet access is available in public areas (surcharges apply). This Region Test property has 50,000 square feet of event space consisting of a conference center, banquet facilities, conference/meeting rooms, and a ballroom. The property has a train station pick up service, which is complimentary. Business services, wedding services, limo/town car service, and tour/ticket assistance are available. Guest parking is limited, and available on a limited first come, first served basis (surcharge). Nearby parking is also available for a surcharge. Other property amenities at this Art Deco property include ski in/ski out access, a concierge desk, and a children’s club. Extended parking privileges may be offered to guests after check out (surcharge). This is a smoke free property. Guestrooms. 1,000 air conditioned guestrooms at Expedia Connect Test Hotel 1 feature windows that open and bathrobes. All accommodations have patios. Rooms are individually furnished. Bathrooms feature jetted bathtubs. Televisions have cable channels. Guestrooms are all non smoking. Official United States of America Rating provided by United States of America Tourism Promotion AuthorityUnited States of America Tourism Promotion Authority assigns an official star rating for accommodations in United States of America. This Apartment property is rated 2.5 keys. Notifications and Fees:Clothing is optional at this property. This property caters to gay and/or lesbian guests. Guests must transfer to the property via airplane and boat and must arrange the transfers by contacting the property before travel, using the contact information on the booking confirmation. Children 6 years old and younger are not permitted from February to January. There are no room charges for children 2 years old and younger who occupy the same room as their parents or guardians, using existing bedding. All guests staying in hotel guestrooms must be registered with the hotel. Some facilities may have restricted access. Guest

EQC Test Hotel (PPP) EUR $2134.76 EQC Test Hotel (PPP) EUR is located in Region Test’s midtown area and near the airport. Property Features. EQC Test Hotel (PPP) EUR’s restaurant serves breakfast, dinner, and brunch. Room service is available. A complimentary breakfast is served in the dining room on weekdays. Recreational amenities include 3 freshwater, heated outdoor swimming pools. Also located on site are a golf course, a children’s pool, a health club, and a fitness facility. Wireless Internet access is available in public areas. The property offers a roundtrip airport shuttle (surcharge). Guest parking is complimentary. Additional property amenities include coffee in the lobby. This is a smoke free property. Guestrooms. 175 air conditioned guestrooms at EQC Test Hotel (PPP) EUR feature coffee/tea makers and complimentary newspapers. Bathrooms feature makeup/shaving mirrors, complimentary toiletries, and hair dryers. Wireless Internet access is available. In addition to desks, guestrooms offer direct dial phones. 32/40 inch LCD televisions have satellite channels. Also included are blackout drapes/curtains and clock radios. Guestrooms are all non smoking. Official United States of America Rating provided by Swedish Hotel and Restaurant Association (SHR)Swedish Hotel and Restaurant Association (SHR) assigns an official star rating for accommodations in United States of America. This Houseboat property is rated 4 star Superior. Notifications and Fees:Guests requiring a visa to enter Russia may be able to obtain help from the hotel where they will be staying. Guests should contact the hotel using the contact information on the booking confirmation. Hotels may charge for providing visa assistance even if guests subsequently cancel their reservation. All arrangements including any applicable charges are made directly between guests and the hotel. In addition, guests will also be liable for any applicable cancellation charges. Couples wishing to share a room must provide proof of marriage. Under Vietnamese law, a guest who is not a Vietnamese citizen cannot share a room with a Vietnamese citizen without proof of marriage to each other. Otherwise, a second room must be booked. To arrange required transfers to the property, guests must contact the property before travel using the contact information on the booking confirmation. A mandatory fee for lunch on 9 November is included in the total price displayed Minimum Spring Break check in age is 27 years old. Children 5 years old and younger are not permitted at this property. The Japanese Ministry of Health, Labour, and Welfare requires all international visitors to submit their passport number and nationality when registering at any lodging facility (inns, hotels, motels, etc. ). Additionally, lodging proprietors are required to photocopy passports for all registering guests and keep the photocopy on file. The following mandatory hotel imposed fees are charged and collected by the hotel either at check in or check out. A fee

API PondCare pH Liquid Test Kit $6.99 Aquarium Pharmaceuticals PondCare Wide Range pH Test Kit Quickly and accurately measures the pH of pond water, reading pH levels from 5.0 to 9.0. Makes 160 tests. Prevents the harmful effects of too alkaline/too acidic pond water resulting from minerals found in tap water, high ammonia levels in the pond, rain runoff or overstocking of pond fish. Includes test solution, glass test tube w/ cap, color chart & instructions. pH is the measure of acidity or alkalinity of water. A pH reading of 7.0 is neutral, a reading above 7.0 is alkaline, and a reading lower than 7.0 is acidic. Many factors can significantly alter the pH of pond water. Acid rain will lower the pH to 6.5 or less. If the carbonate hardness (KH) level is low the pH level will swing up and down over a 24-hour period. Water with a high KH will usually have a high pH. Aquatic plants and algae remove carbon dioxide from the water during the day. This causes the pH to rise above pH 8.0 in the day then fall at night. These fluctuations are normal and will not harm pond life. Extremes in pH, however can stress fish and plants. A pH of 7.0 is considered ideal for plants and fish. Many species of aquatic plants thrive at a slightly acid pH while koi and goldfish prefer a slightly alkaline pH. Therefore, an acceptable pH range is 6.8 to 7.6. Product Features: . Performs about 160 tests . Quickly and accurately tests for pH levels from 5. 0 to 9. 0 Item Specifications: Size: Performs 160 tests Directions for use: Read thoroughly before testing. Do not allow Test Solutions to get into aquarium. To remove childproof safety cap: With one hand, push red tab left with thumb while unscrewing cap with free hand. 1. Fill a clean test tube with 5 ml of water to be tested (to the line on the tube). 2. Add five drops of Indicator Solution, holding dropper bottle upside down in a completely vertical position to assure uniformity of drops. 3. Cap the test tube and invert tube several times to mix solution. Do not hold finger over open end of tube, as this may affect the test results. 4. Determine the pH reading by matching the color of the solution against those on the pH Color Chart. The tube should be viewed against the white area beside the color chart. Color comparisons are best made in a well-lit area. The closest match indicates the pH of the water sample. Rinse the test tube with clean water after each use. Compatability: The Pond Care(R) Wide Range pH Test Kit can be used to test the pH in ponds and water gardens and tap water.

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Powerwh Free Fall Windlasses #36, For Boats Up To 36′, 1/2 Line, 1/4 High Test Chain, 775lb. Max Load, 8 12a Draw, 80′/min Speed Up, 250′/min Down $699 Powerwh Free Fall Windlasses #36, For Boats Up To 36′, 1/2″ Line, 1/4″ High Test Chain, 775lb. Max Load, 8 12a Draw, 80′/min Speed Up, 250′/min Down . Designed to automatically drop ground tackle up to six times faster than comparable power-down models, these helm-operated, free-fall windlasses give you remote operation with simple switch activation, eliminating the need to go forward during anchoring. Drive shafts, rope and chain guide systems, and fasteners are stainless steel. Housings are corrosion-resistant marinegrade aluminum with triple-finish nylon coating. Rubberized deck gasket protects your deck. All models handle both rope and chain, with high-test chain and medium-lay nylon rope, such as New England Ropes Premium Three Strand Line required. Style: Horizontal Rode Type: Rope/chain Dimensions: 6-5/16″H x 7″W x 10-9/16″L Case Material: Nylon-coated aluminum Capstan/Gypsy Material: Stainless steel Warranty: Two years on Free-Fall 31; three years on Free-Fall 36/41/46

Powerwh Free Fall Windlasses #46, For Boats Up To 46′, 5/8 Line, 5/16 High Test Chain, 1100lb. Max Load, 14 18a Draw, 70′/min Speed Up, 350′/min Down $799 Powerwh Free Fall Windlasses #46, For Boats Up To 46′, 5/8″ Line, 5/16″ High Test Chain, 1100lb. Max Load, 14 18a Draw, 70′/min Speed Up, 350′/min Down . Designed to automatically drop ground tackle up to six times faster than comparable power-down models, these helm-operated, free-fall windlasses give you remote operation with simple switch activation, eliminating the need to go forward during anchoring. Drive shafts, rope and chain guide systems, and fasteners are stainless steel. Housings are corrosion-resistant marinegrade aluminum with triple-finish nylon coating. Rubberized deck gasket protects your deck. All models handle both rope and chain, with high-test chain and medium-lay nylon rope, such as New England Ropes Premium Three Strand Line required. Style: Horizontal Rode Type: Rope/chain Dimensions: 6-5/16″H x 7″W x 10-9/16″L Case Material: Nylon-coated aluminum Capstan/Gypsy Material: Stainless steel Warranty: Two years on Free-Fall 31; three years on Free-Fall 36/41/46

Powerwh Free Fall Windlasses #41, For Boats Up To 41′, 5/8 Line, 5/16 High Test Chain, 875lb. Max Load, 10 14a Draw, 75′/min Speed Up, 300′/min Down $769 Powerwh Free Fall Windlasses #41, For Boats Up To 41′, 5/8″ Line, 5/16″ High Test Chain, 875lb. Max Load, 10 14a Draw, 75′/min Speed Up, 300′/min Down . Designed to automatically drop ground tackle up to six times faster than comparable power-down models, these helm-operated, free-fall windlasses give you remote operation with simple switch activation, eliminating the need to go forward during anchoring. Drive shafts, rope and chain guide systems, and fasteners are stainless steel. Housings are corrosion-resistant marinegrade aluminum with triple-finish nylon coating. Rubberized deck gasket protects your deck. All models handle both rope and chain, with high-test chain and medium-lay nylon rope, such as New England Ropes Premium Three Strand Line required. Style: Horizontal Rode Type: Rope/chain Dimensions: 6-5/16″H x 7″W x 10-9/16″L Case Material: Nylon-coated aluminum Capstan/Gypsy Material: Stainless steel Warranty: Two years on Free-Fall 31; three years on Free-Fall 36/41/46

EQC Test Hotel D $216.05 EQC Test Hotel D is located in Region Test.

EQC Test Hotel B $143.85 EQC Test Hotel B is located in Region Test.

EQC Test Hotel J $21 EQC Test Hotel J is located in Region Test.

EQC Test Hotel F $364.36 EQC Test Hotel F is located in Region Test.

EQC Test Hotel A $147.67 EQC Test Hotel A is located in Region Test.

EQC Test Hotel H $174.58 EQC Test Hotel H is located in Region Test.

EQC Test Hotel G $44.49 EQC Test Hotel G is located in Region Test.

Farnam Cough Free 1lb $14.99 Herbal powder treats symptoms of heaves, allergies, stable cough and congestion. All-natural, test free, with no fillers. Works fast. Active Ingredients: Elecampane Root, Ginger Root, Fenugreek Seed, Ferrous Sulfate Monohydrate, Sulphur, Dehydrated Kelp, Dicalcium Phosphate, Comfrey Root, Charcoal, Calcium Carbonate Dosage & Administration: Feed two scoops per day for the first five days. And one scoop a day until coughing discontinues. If the horse isn’t eating well, take 2 scoops in a little warm water and dose syringe into horses throat. Enclosed scoop holds 1/2 oz 1 lb. – 32 day supply (based on maintenance dose) 3 lbs. – 96 day supply (based on maintenance dose) Additional Information Guaranteed Analysis: Protein, min. – 10% Fat, min. – 10% Fiber, max. – 10% Calcium, max. – 1.9% Calcium, max. – 2.4% Phosphorous, min. – 0.8% Iron, min. – 9000 ppm Sulfur, min. – 1%

Farnam Cough Free 3lb $38.99 Herbal powder treats symptoms of heaves, allergies, stable cough and congestion. All-natural, test free, with no fillers. Works fast. Active Ingredients: Elecampane Root, Ginger Root, Fenugreek Seed, Ferrous Sulfate Monohydrate, Sulphur, Dehydrated Kelp, Dicalcium Phosphate, Comfrey Root, Charcoal, Calcium Carbonate Dosage & Administration: Feed two scoops per day for the first five days. And one scoop a day until coughing discontinues. If the horse isn’t eating well, take 2 scoops in a little warm water and dose syringe into horses throat. Enclosed scoop holds 1/2 oz 1 lb. – 32 day supply (based on maintenance dose) 3 lbs. – 96 day supply (based on maintenance dose) Additional Information Guaranteed Analysis: Protein, min. – 10% Fat, min. – 10% Fiber, max. – 10% Calcium, max. – 1.9% Calcium, max. – 2.4% Phosphorous, min. – 0.8% Iron, min. – 9000 ppm Sulfur, min. – 1%

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Weed Free Garden Watering Blanket Kit $79.99 Everything You Need To Start a VegetableGarden Landscape Fabric with Built-in Drip Irrigation! Our Weed Free Watering Blanket offers a new approach to weed blocking and watering – combining two labor and cost intensive steps into one. Now you can enjoy a virtually weed free experience without herbicides. What is Included: 8×10 Garden Blanket 5 lbs of Granular Organic Fertilizer 1 Pint Liquid Organic Fertilizer Soil Test Kit 15 Biodegradable Fabric Stakes Header Hose (pro quality components) 10 PSI Pressure Reducer Benefits of the Weed Blanket: Made with Rainweave WATER WEED & FEED in 1 Step Conserves over 75% on water usage Blocks weeds without chemicals Saves valuable time Increases plant health Organic growing method Increases crop yield Cleaner crops Lasts multiple seasons (5+ years uncovered or 20+ years if mulched)

Dual Sell PDP Test Hotel $227.27 Dual Sell PDP Test Hotel is located in Region Test.

Dual Net OBP Test Hotel $222.22 Dual Net OBP Test Hotel is located in Region Test.

Agency Sell OBP Test Hotel $112.11 Agency Sell OBP Test Hotel is located in Region Test.

Dual Sell PPP Test Hotel $801 Dual Sell PPP Test Hotel is located in Region Test.

Dual Net PDP Test Hotel $278.78 Dual Net PDP Test Hotel is located in Region Test.

Dual Sell OBP Test Hotel $334.33 Dual Sell OBP Test Hotel is located in Region Test.

Dual Net PPP Test Hotel $223.22 Dual Net PPP Test Hotel is located in Region Test.

Audix Tm1 Test And Measurement Mic $299 Audix TM1 Test and Measurement Mic

Rolls Mo2020 Test Tone Oscillator $79.99 Rolls MO2020 Test Tone Oscillator

Shimano Curado 300e Reel, 6.2:1 Gear Ratio, 190/14lb Yds Test, 10.5oz. $249.99 Shimano Curado 300e Reel, 6.2:1 Gear Ratio, 190/14lb Yds Test, 10.5oz. . Everything a high performace, low profile baitcaster aspires to be. Sculpted from aircraft-grade aluminum, its ergonomic frame houses a wish list full of features, including A-RB (Anti-Rust Bearings), Super Free (SF), a Titanium Levelwind Insert and more. It’s a beauty to hold, and a pleasure to fish.br>Features Aluminum Frame Lightweight Graphite Sideplates Recessed Reel Foot High Efficiency Gearing (HEG) Super Free Bearing Supported Pinion Gear System (SF) A7075 Aluminum Magnumlite Spool Dartainium Drag Bearings 4 Shielded Stainless Steel BB Roller Clutch Bearing Variable Brake System (VBS) with Reduced Mass Hub Super Stopper Assist Stopper Septon PV Power Grips 1/8 Turn Easy Access Attached Sideplate Metal Cast Control Knob Anodized Spool and Handle Disengaging Levelwind System Quickfire II Clutch Bar Specifications 12/240, 14/190, 20/120 Test/Yd, 50/205,65/120,80/105 Mono Line Capacity, Right Retreive 15lb. Ma