Demonstration of dimethylnonanoyl-CoA thioesterase activity in rat liver peroxisomes followed by purification and molecular cloning of the thioesterase involved.

Peroxisomes play an indispensable role in cellular fatty acid oxidation in higher eukaryotes by catalyzing the chain shortening of a distinct set of fatty acids and fatty acid derivatives including pristanic acid (2,6,10,14-tetramethylpentadecanoic acid). Earlier studies have shown that pristanic acid undergoes three cycles of beta-oxidation in peroxisomes to produce 4,8-dimethylnonanoyl-CoA (DMN-CoA) which is then transported to the mitochondria for full oxidation to CO(2) and H(2)O. In principle, this can be done via two different mechanisms in which DMN-CoA is either converted into the corresponding carnitine ester or hydrolyzed to 4,8-dimethylnonanoic acid plus CoASH. The latter pathway can only be operational if peroxisomes contain 4,8-dimethylnonanoyl-CoA thioesterase activity. In this paper we show that rat liver peroxisomes indeed contain 4,8-dimethylnonanoyl-CoA thioesterase activity. We have partially purified the enzyme involved from peroxisomes and identified the protein as the rat ortholog of a known human thioesterase using MALDI-TOF mass spectrometry in combination with the rat EST database. Heterologous expression studies in Escherichia coli established that the enzyme hydrolyzes not only DMN-CoA but also other branched-chain acyl-CoAs as well as straight-chain acyl-CoA-esters. Our data provide convincing evidence for the existence of the second pathway of acyl-CoA transport from peroxisomes to mitochondria by hydrolysis of the CoA-ester in peroxisomes followed by transport of the free acid to mitochondria, reactivation to its CoA-ester, and oxidation to CO(2) and H(2)O. (c)2002 Elsevier Science.

Field evaluation of a dog owner, participation-based, bait delivery system for the oral immunization of dogs against rabies in Tunisia.

We evaluated a dog owner, participation-based, bait delivery system for the oral immunization of dogs against rabies. In a field study in a semirural area of northern Tunisia, dog owners were asked to come to temporary bait delivery sites. A total of 314 baits were given to 178 dog owners in four sites. The experimental baits used consisted of a freeze-dried core unit containing sulfadimethoxine (SDM) as a biological marker and an aromatized paraffin envelope. No vaccine was used. Preliminary tests had shown that by using a rapid commercial card test, positive SDM serum levels were detected in more than 95% of dogs up to two days after bait ingestion. During the two days following bait delivery, we visited more than 95% of all households in the study area and took blood samples from as many owned dogs as possible. Unconsumed baits were recovered and human contacts with the bait matrix were recorded. The campaign required 7.6 person-min per bait and 13.5 person-min per dog owner for providing baits, gloves, and instructions. The estimated average cost effectiveness ratio per dog accepting a bait was 1.7 US dollars. From the indications given by the dog owners and the results of the SDM test, it was concluded that 85-90% of the owned dogs in the study area had consumed a bait at least partially. Of 314 baits delivered, 78.7% were fully consumed by dogs and 4.1% were recovered during the household survey. The remaining baits (17.2%) that were not recovered were either not consumed or only partially consumed by the target dogs (3.7 baits per 100 inhabitants). These baits probably remained within the highly populated areas and were potentially accessible to other domestic animals and other nontarget species, including humans. Twenty-five unprotected human contacts with baits were recorded (1.7% of all inhabitants). Our study has demonstrated the potential of dog owner based bait delivery. This technique is simple and efficient, particularly if the human population is accustomed to mass immunization in defined centers. Before applying this method on a large scale with live vaccine loaded baits, further studies should focus on minimizing the number of human contacts with the vaccine bait, systematizing contact identification and establishing structures in ensuring proper treatment if exposure to vaccine should occur.

Field evaluation of two bait delivery systems for the oral immunization of dogs against rabies in Tunisia.

Two bait delivery systems for the oral immunization of dogs against rabies were tested in small scale field trials in a semi-rural area in Tunisia: bait delivery to owned dogs during door to door visits of households (door to door baiting) and distribution of baits on transect lines (transect line baiting). A prototype bait (DBL2) configured for industrial production and containing either sulfadimethoxine (SDM) as a systemic marker or Rhodamine B as a topical marker was used. The overall proportion of dogs which took a bait and presented topical marker staining after door to door baiting was 59.1%. The total time and costs spent per bait accepting dog averaged 34 person minutes and US$4, respectively. Unconsummated baits were readily recovered. No unprotected human contacts with baits were recorded. Door to door baiting is a very specific but time-consuming method that enables a safe administration of vaccine baits to owned dogs. For transect line baiting, baits were distributed at a density of ca 3000 baits per km2 along double transect lines. Baits were recovered after 20 h. According to the proportion of SDM positive serum samples, 24.1% of owned dogs in the baiting area had consumed baits. Of all owned and ownerless dogs, presumably free-roaming during transect line baiting, > 40% had consumed baits. The total time and costs spent per bait accepting dog averaged 48 person minutes and ca US$20, respectively. The household census revealed 32 direct human contacts with the bait matrix which corresponds to 1.4% of inhabitants. Placing baits on transect lines gives the possibility to vaccinate dogs not accessible by vaccination systems which base on dog owner participation. However, the method is not specific, less safe than other systems, not easily accepted by the human population, and costly.

Test of three bait types for oral immunization of dogs against rabies in Tunisia.

Chicken heads and two types of artificial bait were tested in Tunisia during two field trials in a waste disposal site carried out in 1988 and 1989 to compare their effectiveness as vehicles for the oral administration of antirabies vaccine to free-roaming dogs. Baits were made available for 36 hr and those that disappeared or were consumed were replaced on several occasions. In 1988, an artificial bait composed of fat and fishmeal (artificial bait type I) was tested. In the second trial, chicken heads and an artificial bait composed of polymerized fishmeal and wax (artificial bait type II) were compared. The vaccine containers were loaded with a topical marker (rhodamine B or methylene blue) to identify animals that had consumed baits. The artificial type I bait tested in 1988 was poorly accepted, but in the second trial, the number of chicken-head baits probably taken by dogs was more than seven times greater than the number of artificial type II baits taken. Thirteen dogs observed during the day showed topical marker staining. In both trials, most baits were taken during the night when dog activity in the waste disposal site was at its maximum. Artificial baits were characterized either by their lack of thermostability (type I, melting) or a certain attractiveness for cats (type II, fish flavor). Chicken heads fulfill established requirements for baits for vaccine delivery. They are well-accepted by free-roaming dogs, inexpensive, usually easily available at local markets, unattractive to humans, relatively easy to store in large quantities, and easy to handle.