Figure 1.
The bacterial peptidoglycan biosynthesis pathway and antibiotics that interfere with it.
Starting with the transfer of an enolpyruvate to UDP-GlcNAc by the enzyme MurA, shown beside the pathway, MurB catalyzes the formation of UDP-MurNAc in the cytoplasm. MurC, D, E, and F mediate the formation of UDP-MurNAc- pentapeptide. D-alanyl-D-alanine is synthesized by Ddl. Two enzymes, MraY and MurG, form GlcNAc-MurNAc-pentapeptide-pyrophosphoryl-undecaprenol, which is transferred to the periplasm. This disaccharide pentapeptide monomer unit is cross-linked to pre-existing peptidoglycan by penicillin-binding proteins (PBPs) to form the sacculus surrounding the bacterial cell membrane.
Figure 2.
Effects of antibiotics on Closterium cells.
Addition of colchicine led to production of tadpole-shaped cells by inhibiting cell elongation after division. These cells were easy to distinguish from normal crescent-shaped cells. Without antibiotics (Control), cells had two chloroplasts. When Closterium cells were treated with 100 µM ampicillin (Amp), 100 µM D-cycloserine (D-cyc), or 250 µM fosfomycin (FM), cells with only one chloroplast appeared. On the other hand, 100 µM bacitracin (Baci) and vancomycin (VM) did not affect the number of chloroplasts.
Figure 3.
Electron micrographs of ampicillin-treated Closterium cells and of untreated controls.
The magnification in the lower photos is the same as that in the upper photos. Pyrenoids surrounded by starch are indicated by asterisks. Black and white triangles indicate the outer and inner envelopes of chloroplasts, respectively.
Figure 4.
Representative micrographs of ampicillin-treated cells.
For controls, dividing cells in the medium without ampicillin are shown (a1–3). When crescent-shaped cells (a1) enter the division process, chloroplasts started to divide (a2). After septum formation, two new semicells with two chloroplasts appeared (a3). In the medium with 100 µM ampicillin (b1–6), various types of cells were observed. There were normal crescent-shaped cells with two chloroplasts (b1), dividing cells without chloroplast division (b2), divided semicells with only one chloroplast (b3), crescent-shaped cells with one chloroplast (b4), long cells with irregular chloroplasts (b5) and dead cells (b6). The magnification in those photos without a scale bar is the same as that in a1.
Figure 5.
Effects of ampicillin on Closterium cell growth and division.
(A) Growth curves of untreated cells (triangle) and of 100 µM ampicillin-treated cells (square). Measurements were performed in duplicate. (B) Changes in the frequencies of the four cell types: cells with two chloroplasts (Blue), cells with one chloroplast (Red), elongated cells with irregular chloroplasts (Green) and dead cells (purple). Measurements for untreated cells (Cont) at zero and five days and for ampicillin-treated cells (+Amp) are shown.
Figure 6.
Morphological changes in Closterium cells treated with antibiotics.
Without ampicillin (left panel), each of two chloroplasts divides before septum formation. After cell division, the daughter cell restores the crescent-shaped morphology with expansion of the new semicell. Colchicine inhibits elongation of the new semicells at 2 mM. With ampicillin (right panel), cells divide without chloroplast division to form semicells with only one chloroplast. Because cells can elongate without colchicine, crescent-shaped cells with only one chloroplast form. Then, long cells with several nuclei appear due to inhibition of cell division. During a long cultivation, cells die off.