Microbes, Protists and Fungi

Under a microscope, a methane-producing microbe from a cow’s rumen and the gut bacterium Escherichia coli look much alike: small cells with no nucleus. Their ribosomal RNA says they are about as distant from each other as either is from a yeast.

This lesson asks how biologists sort the small things: bacteria, archaea, protists, fungi, and the viruses that are not cells at all. It builds on what an organism has to do and on the surface and size limits in size and scaling.

Three domains from one molecule

Every cell has a plasma membrane (the lipid boundary of the cell), cytoplasm (the contents inside it), DNA as its genetic material, and ribosomes, the particles that make proteins. A eukaryote is a cell, or an organism made of cells, with a nucleus: a membrane-enclosed compartment holding the DNA. Animals, plants, fungi and protists are eukaryotes. Bacteria and archaea have no nucleus, and for a long time they were lumped together as prokaryotes, the “before the nucleus” cells. The organelles of eukaryotic cells are taken up in the measured eukaryotic cell.

That split rests on an absence, and two cells that both lack a nucleus need not be close relatives.

Carl Woese, at the University of Illinois, looked for a character that every cell has. He chose the RNA of the small ribosomal subunit, present in all cells and doing the same job in each. His laboratory and its collaborators grew microbes, including several methane-producing species (methanogens) and typical bacteria, extracted this RNA, cut it with an enzyme, ribonuclease T1, into short pieces, and catalogued the sequence of each piece. Each pair of organisms then got a similarity score, higher when their catalogues shared more pieces.

Under the old split, methanogens, which look like bacteria, should score closest to other bacteria. They did not. In 1977 Woese and George Fox reported that the methanogens formed their own group, about as distant from typical bacteria as from the cytoplasmic ribosomal RNA of eukaryotes. They called it the archaebacteria, a third primary kingdom. In 1990 Woese, Otto Kandler and Mark Wheelis proposed the three domains now in use: Bacteria, Archaea and Eukarya (spelled Eucarya in their paper).

The comparison showed that appearance does not reveal deep relationships and a shared molecule can. It did not observe any ancestor or show how eukaryotic cells arose. How to read the resulting tree is in reading phylogenetic trees, and the deep history in history of life.

Archaea differ from bacteria in more than RNA. Their membrane lipids are built from branched isoprenoid chains joined to glycerol by ether bonds, where bacteria and eukaryotes use fatty acids joined by ester bonds (see membrane structure and function). Their walls lack peptidoglycan, the mesh of sugars and short peptides that forms most bacterial walls; some methanogens have a different but similar polymer, pseudomurein. And archaea are not simply “extreme bacteria”. Many live in soils, oceans and sediments, and methanogens live in the guts of cattle and humans.

Bacteria: shape, envelope, fission and gene swapping

Bacteria come in three common shapes: cocci (spheres), bacilli (rods) and spirilli (spirals). A cell of E. coli is a rod roughly 1 micrometre (um) wide and 2 um long.

The cell envelope is everything that surrounds the cytoplasm. In 1884 Hans Christian Gram published a stain that sorts bacteria by their envelopes. In the method used today, a smear is stained with crystal violet, treated with iodine, washed with alcohol and counterstained with red safranin. Gram-positive cells, such as Staphylococcus aureus and Bacillus subtilis, have a thick peptidoglycan wall that keeps the crystal violet-iodine complex through the wash, so they stay purple. Gram-negative cells, such as E. coli, have a thin peptidoglycan wall beneath an outer membrane; the wash removes the violet and the safranin turns them pink. Not every bacterium has a wall at all: Mycoplasma species lack peptidoglycan and give no useful Gram reaction.

Bacteria and archaea usually reproduce by binary fission: the cell copies its DNA, grows, and divides into two. The doubling time, or generation time, is the time one round takes. Bacteria also gain genes from other cells, which is called horizontal gene transfer. In transformation a cell takes up DNA from its surroundings. In conjugation DNA passes between two cells in contact. In transduction a bacteriophage, a virus that infects bacteria, carries bacterial DNA from one cell to another. The genetics of all three belong to bacterial and phage genetics, and the experiments that showed DNA is the genetic material are in DNA, the genetic material.

Worked problem: how fast does fission add cells? Assume exponential growth: every cell divides once per doubling time, none die, and food and space never run short. Then

N = N0 x 2^g, where g = elapsed time / doubling time

1. E. coli. In rich medium at 37 degrees C, E. coli in the laboratory typically doubles about every 20 minutes. Start with 100 cells. In 3 hours, g = 180 minutes / 20 minutes = 9 generations. N = 100 x 2^9 = 100 x 512 = 51,200 cells.
2. An archaeon. The Asgard archaeon MK-D1, grown by Hiroyuki Imachi and colleagues in 2020, had a doubling time estimated at about 14 to 25 days. Choose 20 days, inside that range, and start counting only after its 30 to 60 day lag, the period before growth begins. Ten doublings take 10 x 20 days = 200 days. For E. coli, ten doublings take 10 x 20 minutes = 200 minutes.
3. Same growth, different clocks. Either way, 1,000 cells become 1,000 x 2^10 = 1,000 x 1,024 = 1,024,000 cells. The archaeon takes 200 days x 1,440 minutes per day = 288,000 minutes, which is 1,440 times as long.

Organism

E. coli: doubling about every 20 min (rich medium, 37 °C, laboratory) Archaeon MK-D1: doubling every 20 days (chosen inside the estimated 14 to 25 days)

Starting cells N0

100 cells 1,000 cells

Elapsed time 180 min (0 to 400 min)

Generations

Doublings

Cells

The same number of doublings on each clock

In this sketch: exponential growth, every cell divides once per doubling time, none die, food and space never run short, constant doubling time, with the curve drawn smoothly so a count between whole doublings is an average; counting for MK-D1 starts after its lag.

Rule to keep: g = elapsed time / doubling time, and N = N0 × 2^g, so equal numbers of doublings give equal cell numbers; only the clock differs. This is a sketch of the logic, not a prediction for a real culture.

Two cautions: real cultures run out of food, and MK-D1 growth was measured as gene copies by qPCR, not cell counts. Still, generation time is one factor in how fast a population, and the genes in it, can change.

Protists: a word for “everything else”

Protist is an informal name for any eukaryote that is not an animal, a plant or a fungus. It is not a natural group. Protists belong to many separate branches of the eukaryote tree, and their classification keeps changing: a 2019 revision of eukaryote classification by Sina Adl and colleagues dropped Excavata as a formal supergroup. Nor are protists “early” eukaryotes; every living lineage has had as long to evolve as animals have.

Compare four:

• Paramecium swims and sweeps food into an oral groove with cilia, many short hair-like projections that beat together.
• Amoeba crawls and engulfs prey by extending pseudopodia, temporary bulges of cytoplasm.
• Euglena swims with a flagellum, a long whip-like projection, and is mixotrophic: it can photosynthesize in light and absorb organic food when light is poor.
• Plasmodium, the malaria parasite, passes between mosquitoes and vertebrates including humans, and in humans multiplies inside liver and red blood cells.

Even mitochondria are not universal. The gut protist Monocercomonoides has no detectable mitochondrion, and its genome suggests the organelle was lost in that lineage, not that it never had one.

Fungi digest outside the body and partner with roots and algae

Fungi are not plants: they do not photosynthesize, and they are more closely related to animals than to plants. A fungus grows either as single cells, a yeast, or as hyphae, threads of cells, which together form a mycelium, the branching mass you see as mould. Fungal walls contain chitin, a tough polymer also found in insect exoskeletons, along with glucans.

Fungi feed by absorption. Hyphae release enzymes into their surroundings, the enzymes break large molecules into small ones outside the fungus, and the hyphae take up the products. Compare an animal, which takes food into a gut first. Baker’s yeast, Saccharomyces cerevisiae, reproduces asexually by budding, pinching a small daughter cell off a larger one, while a filamentous mycelium spreads by growing at its hyphal tips.

The sexual cycle has three steps that are easy to merge. Plasmogamy joins the cytoplasm of two mating partners. Karyogamy fuses their haploid nuclei into one diploid nucleus, and a haploid nucleus carries one set of chromosomes while a diploid one carries two. Meiosis then produces haploid spores (see chromosomes and meiosis). Plasmogamy is not karyogamy. In many fungi, including mushroom-forming species, each cell keeps two separate haploid nuclei, a dikaryon, for long periods before karyogamy.

Fungi also live in partnerships. Mycorrhizae are associations between plant roots and fungi. In arbuscular mycorrhizae the fungus grows into cells of the root cortex, inside the cell wall but outside the cell membrane, and forms branched exchange structures called arbuscules; the mycelium extends far into the soil, and plant and fungus trade carbon from the plant for mineral nutrients, especially phosphorus, from the fungus. Lichens pair a fungus with a photosynthetic alga or cyanobacterium. Some lichens hold more: in 2016 Toby Spribille and colleagues found basidiomycete yeasts in the outer layer of sampled macrolichens, but a 2019 survey of 339 Appalachian lichen species detected such yeasts in only 2.7 percent. For the ecology, see mutualism, parasitism and coevolution, nutrient cycling and soil, nutrients and partners.

Viruses as the contrast case

A virus is a genome, either DNA or RNA, enclosed in a protein coat called a capsid. Bacteriophage T2, which infects E. coli, has a DNA genome; influenza viruses have RNA genomes. A virus has no ribosomes and no cell structure, and it is copied only inside a host cell, using that cell’s machinery.

This course defines an organism as a living individual made of one or more cells. On that definition a virus is not an organism. That is a choice of definition, not a final verdict on what counts as life. Every bacterium, archaeon, protist and fungus in this lesson has a membrane, cytoplasm, DNA and ribosomes; a virus has a genome and a coat. How animals detect and fight microbes and viruses is in innate immunity.

Design problem

Kiers and colleagues reported in 2011 that, in experiments with the barrel medic Medicago truncatula and arbuscular mycorrhizal fungi, plants allocated more carbohydrate to fungal partners that cooperated better, and fungi passed more nutrient to roots that supplied more carbohydrate. Their full methods were not part of this lesson’s sources. How would you test whether a root system rewards the better fungal partner, and whether a fungus rewards the better root?

One way to do it

Give one root system two fungal partners that differ in how much phosphorus they deliver, one generous and one poor.

Label each resource. Feed the leaves carbon dioxide carrying a carbon isotope and measure how much label reaches each fungus. Give each fungus phosphorus carrying a separate label and measure how much reaches the plant.

Add controls. A no-choice control colonizes plants with only one fungus each, which shows whether a fungus draws more carbon wherever it grows or only when the plant can compare partners. In a second run, swap which fungal species is the generous one, separating the partner’s behaviour from its species identity.

Predictions: if plants reward the better partner, the generous fungus receives more carbon in the mixed plants than the poor one does, beyond any difference in the no-choice plants. If fungi reward the better root, more phosphorus flows to roots that supply more carbon.

Limit the conclusion to the species tested: a result in Medicago and two fungi does not cover all mycorrhizal partnerships.