Microbiology

Interactions Between Lake Sturgeon and Aquatic Microbes:
How Do Stream Microbes Affect Lake Sturgeon During Different Life Stages?

 

Microbiology

 

 

 

Aquatic Microbial Communities and Interactions with Fish Eggs

Many aquatic animals spend their entire life from fertilization through death in a dilute solution of microbes that include Bacteria, viruses and unicellular eukaryotes. In oviparous (egg-laying) fishes, eggs extruded into the water column during spawning activities become a surface and substrate on which bacteria might adhere, subsist, and flourish. While some bacterial survival strategies may be deleterious to developing eggs and early life stages, some may be advantageous. These interactions have existed for considerable periods of time, and survival of fish during critical early ontogenetic stages likely depends on an evolutionary balance between acquired characteristics of fish and the abundance and taxonomic composition and diversity of microbial communities within which fish populations have evolved.

 

 

Where Do Interactions Potentially Occur Between Microbes and Fishes?

Unfertilized Egg

  • Microbial communities exist within the female and eggs may be on the egg surface before spawning.

Fertilized Egg

  • Microbes enter through micropile with sperm and are enclosed in the developing egg.
  • Microbes of small size move into and out of the egg through pores together with oxygen (in) and waste products (out).
  • Microbes adhere to the surface of the egg and  potentially affect rates of gas and waste exchange and may be directly pathogenic.

Hatched Larvae During Period of Endogenous Feeding

  • microbes may enter the body through the gills during period of endogenous feeding

Hatched Larvae During Period of Exogenous Feeding

  • Microbes are ingested from the water along with exogenous food sources.
  • Different food sources  may themselves be sources of microbes.
  • Water communities and food sources vary across streams and among sections of  streams affected by human activities.

Microbes are Expelled with Waste Products from Larval and Juvenile Fishes.

  • The likelihood of intake in the body will be dictated by aquacultural practices or in streams by inputs of microbes and chemicals from outside sources.

 

 

 

 Human Impacts

Different streams are characterized by different physical and  chemical properties that fishes adapt to.  Microbial communities   (a) likewise affected by these properties and (b) are part of the   biotic community to which fish adapt.

Different fish species are characterized by different life histories and different physiological, morphological features that are adapted to conditions in the aquatic environment.  Disruption of communities affect the abilities of fish to survive and to adapt to changing conditions.

Different human stressors including presence of dams, and activities that change the physical and chemical and biological properties of aquatic communities are disrupting linkages and reducing adaptive potential.

 

Water Quality Effects

 

 

General Features of Sturgeon Eggs Colonized by Bacteria

Egg Bacteria

 

 

 

Figure 1. Interactions of microbes with sturgeon eggs.
Panel A. Sturgeon eggs preserved in 80% ethanol.
Panel B. An egg sliced through the middle and then viewed under the microscope focusing on the boxed area would produce the images in
Panel C (100X magnification) &
Panel D (1000X magnification). The interior of the egg is protected from the environment by the tough chorion layers.
In Panel D one can see the outer microfibrous layer of the chorion with bacteria stuck to the outside (arrows). The lines through the chorion are channels for exchange of gases with the environment by the developing embryo. Panels E-H are scanning electron micrographs taken at 5000X magnification of the surface of sturgeon eggs. Panels E and G show the surface of eggs that have been aseptically harvested from female surgeon, hence they have not been in the water and exposed to bacteria. Bacteria are not detectable on the surface.
Panels F & H show the surface of eggs that have been fertilized and exposed to stream water for several days. These eggs are covered with bacteria of many different shapes.

 

 

Percentages of different microbial species present in the stream and on the egg surface at different times.  Different colors represent different taxa of microbes.  Note the different percentages of microbial types in water and on the egg surface during different periods during incubation.

Egg Microbe Species Chart

 

 

 

Phylogenetic composition of egg-associated bacterial community. Two groups of microbes that are known fish pathogens are identified.

Distribution of 16S rRNA microbial taxa

16S Phylotypes

 

 

 

 

 

 

 

PCA Analysis of Collected rRNA DNA Sequences showing differences in microbial community composition in the water (WL and WE for water communities collected late and early in the spawning period, respectively, and on the eggs during different periods of incubation.  The eggs communities are not representative of what is in the water.

 

 

 

PCA Graph

 

Sturgeon – Microbe Experimental Design

Microbe Experimental Design

 

 

 

 

Factors that affect egg mortality:

  • Temperature (warm or cold)
  • Filtration (filtered or non-filtered)

Microbe Experimental Design

Microbe Experimental Design

 

 

 

 

 

 

The effects of water filtration on egg mortality. Sturgeon eggs that were reared in non-filtered water had significantly higher egg mortality than eggs reared in water that had been filtered to remove microbes.

Water Filtration Chart

 

 

Daily Mortality Rates During Incubation by Family

 

Early Spawning Females

Early Spawning Graph

 

Late Spawning Females

 

 

 

 

 

 

 

 

 

 

The length of the incubation period varies as a function of water temperature as shown in the two figures of eggs from ‘early’ and ‘late’ spawning females in cool and warmer water.  The majority of the mortality occurs at the beginning of the second trimester of incubation.  We feel that the abundance and diversity of microbes in the water and on the surface of the egg will determine the levels of mortality experienced.

 

 

 

Effects of Microbes on Larval Phenotype at Hatch

Water Filtration Chart
Hatch Yolk Sac

 

Both water temperature during incubation and filtration affected the amount of yolk sac energy reserves that were present at hatch.  Our interpretation is that eggs incubated in unfiltered water had to use energy reserves while in the egg to mount a defense against microbes, leaving fewer energy reserves available for growth after hatch.

 

 

The Effect of Water Treatment on Yolk Sac Resource Uses

 

 

 

 

Eggs reared in stream water had significantly smaller yolk sac area than that reared in filter treated water.

This suggests that eggs used yolk sac resources for defense against microbes on the egg surface.

 

Yolk Sac Chart

 

 

Conclusions

  1. Sturgeon eggs provide an excellent substratum for bacterial populations
    1. We are now quantifying the number of bacteria per egg as a function of ecosystem variables
  2. The eggs appear to select for specific populations of bacteria
    1. We are now quantifying the number of bacteria per egg as a function of ecosystem variables
    2. Are the selected populations symbionts?
  3. The nature of the selection(s) is unknown
    1. Chemistry of attachment
    2. Excreted waste & possibly specific attractants/repellants
  4. Microbial communities as biotic agents exerting direct (egg survival) and indirect effects (early life history traits of larval fishes)

 

 

Conclusions

  • timing of reproduction varies among adults but in predictable ways
  • covariation of early life history traits with environmental conditions during spawning
  • influence of microbial communities on survival and traits
  • community composition varies over time and specificity to eggs
  • some taxa have known pathogenic effects and others may be beneficial
  • maternal effects – anti-microbial (lysozyme) present in the female and on the egg surface

Conclusions