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In 2001-2002 throughout Thailand, black tiger shrimp Penaeus monodon farmers reported very unusual retarded growth. We have called this problem monodon slow growth syndrome (MSGS). Based on decreased national production, estimated losses due to this phenomenon were in the range of 13 000 million baht (approximately US $ 300 million) in 2002. Since rearing practices had not changed, it was considered possible that the MSGS problem may have arisen from a new or existing pathogen. To examine this possibility, cultivated shrimp were sampled from 32 commercial rearing ponds that reported abnormally slow growth from eastern, central and southern regions of Thailand. Shrimp were randomly sampled from each pond and grouped into normal and small shrimp. Normal shrimp were defined as those with body weights (BW) of 24 g or more while small shrimp were defined as those that weighed 16.8 g or less. Pleopods were used for detection of monodon baculovirus (MBV), heptopancreatic parvovirus (HPV) and infectious hypodermal and hematopoietic necrosis virus (IHHNV) using specific polymerase chain reaction (PCR) assays. In addition, some shrimp were processed for normal histopathology and transmission electron microscopy (TEM). Most of the shrimp specimens were infected by at least 1 of these viruses but many had dual or multiple infections. Prevalence of HPV and combined HPV/MBV infections in the small shrimp was significantly higher than in the normal shrimp. In addition to the viruses, a new microsporidian species, gregarines and bacteria were also observed but were not significantly associated with the MSGS problem. Some of the small shrimp gave negative results for all these pathogens by PCR and histology and no new and unique histopathology was recognized in any of the samples. The findings suggested that HPV infection was a contributing factor but not the overriding factor responsible for MSGS. It is possible that MSGS is caused by an unknown pathogen or by some other presently unknown, non-pathogenic factor. KEY WORDS: Penaeus monodon · Growth retardation · MBV · HPV · IHHNV · Microsporidian · GregarinesResale or republication not permitted without written consent of the publisher

Yellow-head virus (YHV) causes acute infections in Penaeus monodon that result in very high mortahty First reports of the vlrus suggested that the viral core consisted of DNA and that the virus should b e classified as a granulosls-type baculovirus However, 3 attempts at DNA extraction with high concentratlons of punfied virus (verified by transmission electron micioscopy, TEM) gave only traces of DNA, which could not be visuahzed by ethidium bromide staining of agarose electrophoresls gels Although selected recombinant clones denved from these pooled DNA traces did not hybridize with host s h n m p DNA, they also tailed to react with YHV-~nfected tissue by the In s~t u DNA h y b n d~z ation technique Furthermore, negatively stained virions of YHV viewed by TEM were atypical for baculov~ruses and viral assembly IS cytoplasmic Therefore, renewed attempts to extract n u c l e~c acid fiom punfied YHV preparations focused on RNA rather than DNA Hemolymph was collected aseptlcally by syringe from 200 artificially YHV-infected, live s h n m p in terminal stages of the disease Purlfled vlrions were prepared by a program of centrifugation culminating In 22 % to 45 , Urografin gradient ultracentrifugation A band at the 30-37% Interval of the gradient gave the cleanest preparation with the highest quantity of vinons By TEM these were enveloped measured 150-170 X 40-50 nm and were surrounded by a f r~n g e of knob-hke projections approximately 11 nm in length Nucleic acld was extracted using g u a n i d~u m thiocyanate and punfied by CsCl gradient ultracentnfugation High-molecular-weight nucleic acid was obtained which was degraded by RNase-A but not by DNase I Based on morphology of negatively stained virions by TEM and on RNA content YHV resembles rhabdoviruses or coronaviruses, rather than baculoviruses This 1s an important discovery since i t necessitates cDNA preparation in the process to develop a n u c l e~c -a c~d probe for YHV detection by the In sjtu or dot blot hybndization techniques

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