Abstract
Stress is one of the most critical factors in fish health management. pH is an important chemical
stressor which indicates the acidity or alkalinity of the aquatic environment. Now a day's acidification
of open and closed water ecosystem is recognized as a global environmental problem. Zebrafish can
tolerate a wide range of pH i.e. 4.3-10.7. The aim of this study was to examine the acute and chronic
stress effects of pH on some biochemical (blood glucose) and hematologial (hemoglobin, red blood cell
and white blood cell) responses, egg production, embryonic development, heartbeat, hatching rate,
behavioral responses and alternation of different organs (liver and kidney) of zebrafish (Danio rerio).
To determine the stress effects of pH, zebrafish were exposed to sub-lethal levels of acidic (5.0-5.5) and
basic (9.5-10.0) pH for 120 hrs and 90 days for acute and chronic stress responses, respectively. Control
groups were maintained at pH 7.2-7.4. In acute stress responses, blood glucose and hemoglobin level
varied significantly in acidic and basic conditions compared to control pH. A significant reduction in
hemoglobin level and RBC concentration and significant increase in WBC concentrations were
recorded in extremely acidic and basic pH in chronic stress condition. Zebrafish embryos were
exposed to different concentrations of pH (2, 3, 4, 5, 6, 9, 10, 11 and 12) for 120 hrs and determined the
effects on the embryonic development and hatching rates. No eggs hatched out at extremely acidic (2
and 3) and basic (11 and 12) pH. Cleavage rate pattern of embryos were not synchronous at pH 4, 5, 10
and 11. Hatching rates were also reduced significantly at pH 4, 5, 6, 9 and 10 from control pH.
Recorded hatching rates were 25%, 45%, 72%, 60%, 57% and 90% at pH 4, 5, 6, 9, 10 and control,
respectively. The result also revealed that with the increasing of developmental stages, pH tolerance
was also increased. Gastrula stage embryos were more tolerant than blastula stage and 2 to 4 cell stage
embryos. During the early stage of development, pH exposures only for a brief period also exerted
negative effects on the embryonic development and hatching rate of zebrafish. During the embryonic
and larval development, the acidic and basic pH significantly increased the percent of abnormality
compared to control pH. The highest percent of abnormality (40%) in different organs was recorded at
pH 5.2 whereas the percent of abnormality in control pH was only 1.64%. Different developmental
anomalies in different organs were recorded as edema and blood clotting in yolk sac, unstraight
notochord, ruptured dorsal and tail fins, scoliosis, twisted tail, ocular hypertrophy and growth
retardation in acidic and basic pH. Highly acidic (5.2) and basic (10.0) pH also affects the
heartbeat/min of zebrafish. The heartbeat/min was increased significantly in acidic and basic pH
compared to control pH. In chronic stress responses, egg development and egg production of
zebrafish were significantly reduced in acidic and basic pH. The result also showed that basic pH has
more negative impacts on the egg production than acidic pH. When zebrafish were exposed to acidic
and basic pH for long time, they showed some behavioral responses like sluggish movement, high
and abnormal mucus production and opercular movement, slow feeding activity etc. In chronic stress
responses, Zebrafish were exposed to acidic (5.0-5.5) and basic (9.5-10) pH for 15 and 30 days. pH has
caused some histological alternations in the liver and kidney of zebrafish. After 30 days of pH
exposures, severe histological alternations in the liver were recorded as pyknotic nucleus,
melanomacrophages aggregate, nuclear hypertrophy, focal necrosis etc. The results also demonstrated
that the histopathological changes in the kidney of zebrsfish were necrosis, ruptured kidney tubules,
tumor formation and glomerular expansion. In conclusion, the results showed that exposure to both
acidic and basic pH exerts severe stress, which affect on the total physiology of fish and offers a
simple tool to evaluate the potential risk of polluted water caused by pH to fish.