In the field and in the laboratory

The first synchronous liberation event was observed on the fourth night following Full Moon, in April 1985. This suggested a hypothesis in answer to one of the important questions driving the study: is reproduction of Millepora platyphylla periodic? My interest was piqued.

I started a more systematic mode of collection, focusing on Toguan Bay: colonies were numbered by writing on their surfaces with plain graphite pencils. Numbered colonies were visited occassionally, I went to Toguan, and fragments collected from them. Toguan Bay is a leeward reef. I mostly dove at Toguan during the late afternoon or early evening.

The bottom at Toguan drops off rapidly. Millepora platyphylla, as well as the few colonies of Millepora dichotoma, resided mainly in a narrow band close to the reef margin. Corals were studied in the upper 10 or 12 feet. Swimming southward along the reef, most of the colonies could be visited along the way, sequentially, approximating a transect from the Toguan River toward Bile Bay, running probably not more than 200 meters (estimated 23 years later).

Numbered colonies were sampled sporadically, on days when the reef was visited. An attempt was made to visit the reef throughout the monthly cycle, Fragments of various sizes were broken off, marked with the colony number, and site, and dated, in pencil before placing them in large ziploc bags (probably Ziploc Freezer Bags, 1 gallon size). Alternately, each bag had a numbered tag, and the number and colony number written on a slate and recorded back at the lab or on the beach.

Back at the laboratory (the UOG Science Building had a well stocked microscopy laboratory) or the Marine Laboratory, colonies were prepared. Later in 1985, when I was able to utilize the microscope laboratory, many, if not all, specimens were broken, and a small piece placed in a fixative. Numerous fixatives were experimented with: 5-10% formalin in seawater; Bouin's fluid; Mercuric Chloride fixatives; Susa's; Dichromate---a large number. I had purchased a copy of Grays' Microtomist's Formulary and Guide, and I attempted to use as many fixatives and stains as possible.

Hard parts were filed separately, in plastic bags. They are still available in 2008, at the UOG Marine Laboratory. Fixed specimens were moved into various decalicfying agents. I experimented with Formic Acid, reasoning that the shock (or so I imagined) to the tissues/cells would be less, if the same basic functional groups were present. Bouin's Fluid has a decalicifying nature, and several changes were often made. A number of other agents were utilized. EDTA was used briefly, a time or two.

In decalicifying agents, the tissues readily separated from the Calcium Carbonate skeleton, floating up or resting on the dissolving skeleton, in a matter or hours or days, depending on the agent and its strength. These sheets were easily trimmed with scissors, and moved through a series of solutions, in the process of embedding them in wax blocks. They are little thicker than a sheet of tissue paper.

I also attempted to observe the living polyps in the stereomicroscope. Duerden had observed Millepora spp. in Jamaica, near the turn of the 19th to the 20th Century. He found it impossible to observe them in the laboratory, when he took freshly broken fragments into the laboratory from the reef: they would not open up. His solution was to break fragments off and leave them on the reef, for several days, or longer, allowing them time to heal.

I did not repeat Duerden's experiments. I attempted several times to see open polyps of Millepora platyphylla. Once I was able to do so. Only once. I observed a gastrozoid open. I applied intra vitam Methylene BLue, and observed the nerve ring located around the margin.

Next: Microscopical studies.