Coliform Bacteria: Occurrence and Antibiotic Susceptibility

Coliform Bacteria: Occurrence and Antibiotic Susceptibility Abstract   Bacteriological contamination of muscles and digestive tract contents of Oreochromis sp. and Labeo sp. reared in a pond supplied with domestic sewage was enumerated followed by determination of resistance of thermotolerent coliforms for antibiotics. Numbers of bacteria in muscles and digestive tract contents of fish reflected their densities in water. Muscles of both the fish species contained high numbers of total coliforms (TC) and fecal coliforms (FC). Escherichia Coli (E.coli) were never recovered from the muscles but from the digestive tract contents of the fish. Ranking of the total and fecal coliform contamination levels showed a decrease in the order digestive tract contents> muscles (p Keywords: Domestic sewage; Fish; Fecal coliforms; Escherichia coli; Water quality; Antibacterial resistance; Public health. Introduction All around the world, people both in rural and urban areas have been using domestic wastes to fertilize fish ponds (Strauss et al. 2000). In the majority of cases, domestic sewages are applied untreated or only partially treated through storage (Strauss 2000). Domestic wastewater, rich in nutrients, used in aquaculture supports the growth of plankton and other microorganisms which are consumed by the fish with little intake of other supplemented feed. Recycling of domestic sewage through aquaculture is an effective form of pollution control, which contributes to cost recovery and provides a source of low cost animal protein production. Domestic sewage transports a variety of human pathogenic microorganisms which may contaminate fish flesh when fish is grown in ponds receiving waste water (Niewolak and Tucholski 2000). Besides, municipal wastewater consists of huge amounts of incompletely metabolized antimicrobial drugs which can lead to the development of antibiotic resistant bacteria as well as resistant plasmids (Wiggins et al. 1999). The prominently affected bacteria are members of enterobacteriaceae and related gram negative rods (Kelch and Lee 1978). One of the important concern of wastewater fisheries is the contamination of fishes by fecal coliforms (Fapohunda, MacMillan, Marshall and Waites 1994). Their presence in fish intended for human consumption may constitute a potential danger not only by causing disease but also because of the possible transfer of antibiotic resistance from aquatic bacteria to human-infecting bacteria from nonaquatic sources (Olayemi, Adedayo and Ojo 1991). Therefore, periodic and c omprehensive sanitary survey of wastewater fishery is required. For years, the group of fecal (also called thermotolerant) coliforms (FC) has been the most widely used as fecal contamination as their excreted load is similar or larger than that of pathogenic organisms, and their survival time in the environment longer than that of excreted bacteria and viruses (Strauss 1997). In the present study, an attempt has been made to determine the bacteriological contamination of muscles and digestive tract contents of Oreochromis sp. and Labeo sp. reared in wastewater fed pond. Resistance to two very common antibiotics for random thermotolerent coliform isolates from muscle and digestive tract contents of both the fish was also determined. Materials and Methods Study Site A sewage fed pond of Bandipur, Rahara, North 24 Parganas, (22 °44N Latitude and 88 °24E Longitude) was taken into consideration for this study and to examine bacterial load of water and fish. Raw sewage was entirely of domestic origin, coming from Titagarh town of North 24 Parganas, West Bengal. Sampling and Dissection Fish samples were caught with a net and were immediately transferred to the laboratory in containers with pond water. They were dissected according to Buras et al. 1987. Muscles and digestive tract contents were isolated and placed in sterile glass vessels. The tissues were weighed under sterile conditions, ground in a mortar and suspended in sodium chloride (NaCl) physiological solution (10 ml of the solution for each 1 g of the muscle or digestive tract content). The suspensions were homogenized using Universal Laboratory Aid Type MPW-309 homogenizer, at 1000 rpm, for 10 minutes. The homogenates were then serially diluted (10-1 to 10-6 for muscles and 10-1 to 10-7 for digestive tract contents) and inoculated into culture media. Time lag from fish collection to the analyses did not exceed 6 hours. Water from sewage-supplied pond was sampled and analysed simultaneously with fish sampling. Samples were collected monthly from July 2009 to September 2009. Microbiological Analyses Total Coliforms Lauryl Tryptose (LT) Broth at 350C for 48 hr was used for three-tube most-probable-number (MPN) presumptive determinations of coliforms (APHA 1998). From all positive presumptive tubes, total coliforms were confirmed by the formation of gas in any amount in the Durham fermentation tubes of brilliant green lactose bile broth (BGLB) for 48 hr at 350C. Fecal Coliforms and E.coli All positive Lauryl Tryptose (LT) MPN tubes to tubes of Escherichia coli (EC) Broth followed by incubation at 44.50C for 48 hr constitute a positive fecal coliform test. The growth from positive EC tubes was then streaked onto Levine Eosin Methylene Blue (EMB) Agar plates and incubated at 35 °C for 18 to 24 h. Colonies from EMB Agar plates typical of E. coli were transferred to Nutrient agar (NA) slants from which GIMViC tests were performed where G-medium is the secondary EC broth, I -medium is Tryptone broth, M- and V-medium is Buffered Glucose broth, and C-medium is Simmons Citrate agar. MPN of E. coli was then computed based on the number of tubes found to contain isolates that produce GIMViC reaction patterns characteristic of E. coli (APHA 2001). Representatives of typical thermotolerent coliform isolates from fish samples were selected randomly by colony morphology on Eosin methylene blue agar and were streaked aseptically several times on freshly prepared nutrient agar plates to obtain pure isolates (Ogbonna, Sokari and Amaku 2008). Nutrient agar plates were then supplemented with ampicillin (50 µg ml-1) and tetracycline (25 µgml-1) and were used to evaluate antibiotic susceptibility patterns of 117 pure isolates (Miranda and Zemelman 2001). 32 isolates from muscles and 24 isolates from digestive tract contents of Oreochromis sp. and 39 strains from muscles and 22 isolates from digestive tract contents of Labeo sp. were subjected to antibiotics sensitivity test. Statistical Analyses Means and standard errors (SE) were calculated. T test was performed between bacterial concentration of muscles and digestive tract contents of both the fish. A significance level of 5% was considered (Zar 2007). Results Bacterial loads in muscles and digestive tract contents of Oreochromis sp. and Labeo sp. were exceptionally high. Total coliforms and fecal coliforms were commonly found in all analysed fish tissues. Escherichia Coli were not found in the muscles of either fish. Additionally, however, thermotolerant Escherichia coli were present in the digestive tract contents of both the fish (Table 2). Bacterial loads in the fish were significantly higher (p Antibiotic resistance pattern Of the 117 thermotolerent coliform isolates examined for antibiotic sensitivity, 82% (96 isolates out of 117) were tetracycline resistant and 65% (76 isolates out of 117) were ampicillin resistant. 58.11% (68 isolates out of 117) of the total isolates were resistant to both antibiotics where as 31% (36 isolates out of 117) were resistant to single antibiotic. From the single antibiotic resistant isolates, 24% were tetracycline resistant and 7% were ampicillin resistant (Figure 1). 56.41% isolates (22 isolates out of 39) from flesh and 22.72% (5 isolates out of 22) from digestive tract contents of Labeo sp. showed resistance to both antibiotics whereas 71.87% isolates from flesh (23 isolates out of 32) and 75% (18 isolates out of 24) from digestive tract contents of Oreochromis sp. showed resistance to both ampicillin and tetracycline (Figure 2). Discussion Comparison of fecal coliform counts of water of Bandipur sewage fed fish pond with WHO (World Health Organization) water quality criteria (WHO 1989) suggests considerable contamination of the first. Bacterial flora of fish reflects the bacteriological quality of the water from where the fish harvested (Geldrich and Clarke 1966). Strong correlation between the bacterial species present in the pond water and the fish regardless of the type of fish were also reported by Buras et al. 1987; Ogbondeminu 1993; Apun, Yusofand and Jugang 1999. Thus, in our study, total coliforms, fecal coliforms and E.coli recovered from muscles and digestive tract contents of Oreochromis sp. and Labeo sp. may reflect bacteriological water quality of the Bandipur sewage fed pond. Fecal coliforms in fish muscles were recovered when values of FC in water were 3.86+3.63105 MPN 100 ml-1 which were much higher than those recommended by WHO (1989) in its health guidelines on wastewater use in aquaculture. Fecal coliforms in fish reflect the level of pollution of their environment, as the normal floras of fish do not include coliforms (Cohen and Shuval 1973). Presence of fecal coliforms indicates the presence of fecal material from warm-blooded animals. However, thermotolerent coliforms include the genera of fecal as well as non fecal origin. E. coli is a species of fecal coliform bacteria that is specific to fecal material from humans and other warm-blooded animals (Bhatia 2008). Environmental Protection Agency (1992) thus recommends E. coli as the best fecal indicator of health risk from water. No detectable penetration of E.coli in muscles of either fish was found at 1.34+0.95104 MPN 100 ml-1 of E.coli concentration in water of sewage fed pond (Table 1). Thus, th e fish flesh qualities at harvest were good on the basis of their E. coli counts. Safety precautions during fish processing are still needed to avoid cross-contamination due to high accumulation of microorganisms in the digestive tract of fish. In this study significantly higher numbers of bacteria (p A wide range of thermotolerent coliforms isolated from sewage fed fish showed resistance to both ampicillin and tetracycline. Multiple anitibiotic resistant faecal coliforms have been observed in wastewater across the world (Gallert et al. 2005). Antibiotic resistance among random bacterial isolates from different organs of fish captured from fecally contaminated water with a full range of resistance (00-100%) to different common antibiotics of therapeutic and prophylactic use among human beings and in various animal farms and fish farms was reported by several authors. (Rhodes et al. 2000; Miranda and Zemelman 2001; Pathak and Gopal 2005). Thus the source of the problem of antibiotic resistance bacteria in wastewater pond of Bandipur was fecally contaminated water. Among thermotolerent coliforms recovered from fish, resistance to ampicillin and tetracycline was found in 65% and 82% of the isolates, respectively. Occurrence of thermotolerent coliforms with high resistance to ampicillin and tetracycline reflect human influence in the environment (Andersen and Sandaa 1994). Domestic sewage enters into the pond environment of the sewage fed farm with huge antibiotics which are used as medicines, as growth promoters or as preventative maintenance and may have established a selective pressure due to a slow degradation of antimicrobials favouring further growth of antibiotic-resistant bacteria (Petersen and Dalsgaard 2003). It may possible that these antibiotic resistant bacteria from wastewater may transfer their antibiotic resistant determinants to indigenous flora of fish, provoking their spread and prevalence in aquatic environment. In the present study bacteria resistant to both ampicillin and tetracycline from digestive tract contents were higher in Oreochromis sp. than in Labeo sp. It may be related to detritus feeding habit of Oreochromis sp. by which it is more exposed to wastes as well as antimicrobials than Labeo sp. which is a column feeder. Similar findings were reported by Miranda and Zemelman (2001) with demersal and pelagic fish. Antibiotic resistant fecal bacteria form domestic sewage may change nutritionally beneficial intestinal microflora with unexpected consequences on fish health. Our study indicates that fish flesh qualities were satisfactory in terms of E.coli counts. In spite of that flesh of both fish showed high numbers of antibiotic resistant thermotolerent coliforms which may include Klebsiella spp., Citrobactor spp. and Enterobacter spp. (non fecal origin) but till have immense ecological and public health implications specially if the resistance is plasmid mediated then there could be a problem associated with the transfer of resistance determinants to human pathogenic bacteria which may enter in human population through fish consumption. According to Walia et al. (2004) antibiotic resistance genes against ampicillin, streptomycin, and tetracycline are known to be transferable to other bacteria. Thus, we can say that Wastewaters and fishes reside there are potent source of antibiotic-resistant bacteria, which in turn may transfer their resistance genes to nonresistant bacteria (Schwartz et al. 2003). Several studies indicate that the environmental conditions in wastewater may enhance the likelihood of gene transfer (Pote et al. 2003). Mach and Grimes (1982) demonstrated the high transfer frequencies of enteric bacteria in a wastewater. Additionally resistant bacteria may pose a risk of therapeutic problems to public health and fish population. So the study demands an elaborate investigation on the members of predominant multidrug resistant bacterial microflora associated with sewage fed fishery along with their plasmids profile as an evidence of conjugal transfer of antibiotic resistance genes in human and animal food chain through fish consumption.

Boeing:how low can they fly? Essay

Prepare problems 3, 5 and 7 on pp.521 and the additional problem, Principles of Economics, Case, Fair and Oster. Problem 3 p.521 For each of the following statements, decide wether you agree or disagree and explain your answer : 1. During period of budget surplus (when ? < ? ), the government debt grows. 2. A tax cut will increase the equilibrium level of GDP if the budget is in deï ¬ cit but will decrease the equilibriumm level of GDP if the budget is in surplus. 3. If the ? ? ? = 0.90, the tax multiplier is actually larger than the expenditure multiplier. Problem 5 p.521 Expert economists in the economy of Yuk estimate the following : Real output/income 1000 billion Yuks Government purchases 200 billion Yuks Total net taxes 200 billion Yuks Investment spending (planned) 100 billion Yuks Assume that Yukers consume 75 percent of their disposable incomes and save 25 percent. 1. You are asked by the business editor of the ? to predict the events of the next few months. By using the data given, make a forecast. (Assume that investment is constant.) 2. If no changes were made, at what level of GDP (? ) would the economy of Yuk settle ? 3. Some local conservatives blame Yuk’s problems on the size of the government sector. They suggest cutting government purchases by 25 billion Yuks. What effect would such cuts have on the economy ? (Be speciï ¬ c) 1 Problem 7 p.521 Assume that in 2008, the following prevails in the Republic of Nurd : Y=200$ C=160$ S=40$ I(planned)=30$ G=0$ T=0$ Assume that households consume 80% of their income, they save 2% of their income, ? ? ? = 0.8 and ? ? ? = 0.2. That is, ? = 0.8 and ? = 0.2 . 1. Is the economy of Nurd in equilibrium ? What is Nurd’s equilibrium level of income ? What is likely to happen in the coming months if the government takes no action ? 2. If 200$ is the  « full-employment  » level of ? , what ï ¬ scal policy might the government follow if its goal is full employment ? 3. If the full-employment level of ? is 250$, what ï ¬ scal policy might the government follow ? 4. Suppose ? = 200$, ? = 160$, ? = 40$ and ? = 40$. Is Nurd’s economy in equilibrium ? 5. Starting with the situation in part (4), suppose the government starts spending 30$ each year with no taxation and continues to spend 30$ every period. If ? remains constant, what will happen to the equilibrium level of Nurd’s domestic product (? ) ? What will the new level of ? and ? be ? 6. Starting with the situation in part (4), suppose the government starts taxing the population 30$ each year without spending anything and continues to tax at that rate every period. If ? remains constant, what will happen to the equilibrium level of Nurd’s domestic product (? ) ? What will the new level of ? and ? be ?. How does your answer to this question differ from your answer to question (5) ? Why ? 2 Additional problem INCOME 2000 3000 4000 5000 6000 7000 8000 9000 10000 TABLE 1 – Add caption TAX C G I 200 1400 1600 1000 200 1900 1600 1000 200 2400 1600 1000 200 2900 1600 1000 200 3400 1600 1000 200 3900 1600 1000 200 4400 1600 1000 200 4900 1600 1000 200 5400 1600 1000 ? 1. Fill in the disposable income ( ) and the saving (?) columns. 2. Deï ¬ ne  « equilibrium income/output  » and then ï ¬ nd the equilibrium income / output by two different methods. 3. Deï ¬ ne ? ? ? and ? ? ?, and ï ¬ nd their numerical value. 4. How much is the : – Government expenditure multiplier ? What is the formula ? – Consumption multiplier ? What is the formula ? – Investment multiplier ? What is the formula ? – Tax multiplier ? What is the formula ? 5. From the above data : – Derive the consumption equation – Derive the saving equation – Derive the investment equation – Derive the government expenditure equation 6. What will be the new equilibrium income when the government increases its expenditure from 1600 to 3100 ? 7. What will be the new equilibrium income when the government increases the taxes from 200 to 1200 ? 8. Assume that the equilibrium level on income/output is the one found in question (2) above. If the potential income/output is 9000 and the government intends to bring the economy to the potential level : – By how much government expenditures should increase/decrease ? Explain. – By how much taxes should increase/decrease ? Explain. 3

How to Choose Tkam Argumentative Essay Topics

How to Choose Tkam Argumentative Essay Topics There are just a few things that define whether an essay you're working on is going to be a good one. It is possible to also restate the ideas you've discussed in the body paragraphs in order to make your point valid. After you settle on the subject and pick the position on which you will base your essay, the remainder of the job can then begin. Nonetheless, the procedure will certainly help you gather your thoughts, and you'll eventually find something worthy of writing on. What Is So Fascinating About Tkam Argumentative Essay Topics? Understanding how to compose a strong argumentative paper can help you advance your very own argumentative thinking. Bear in mind you may make funny argumentative essays if you do a few things. When you're picking your topic, remember that it's much simpler to write about something which you presently have interest ineven in case you don't know a great deal about it. 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Alexander The Great, King Of Asia - 2104 Words

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The Cask Of Amontillado By Edgar Allan Poe - 707 Words

In the short story of The Cask of Amontillado, Edgar Allan Poe writes in first person point of view from the perspective of Montresor who seeks revenge against Fortunato. Montresor began to develop the perfect plan for revenge. During the carnival season, Montresor meets with Fortunato and decides to implement his plan carefully through irony. Poe s story describes the murderer s mind which has lived as a memory of Fortunato s death for fifty years. Poe uses different types of irony and symbolism in the conversations between Montresor and Fortunato which are discussed in the following paragraphs. First, Poe uses dramatic irony in the story. For example, Montresor expresses concern about Fortunato s and says, Come, I said, with decision, we will go back; your health is precious. You are rich, respected, admired, and beloved; you are happy as once I was. You are a man to be missed. For me it is no matter. We will go back; you will be ill, and I cannot be responsible (Poe). 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The purpose is to get the reader to no longer be the observer. He wants them to see with MontressorRead MoreThe Cask of Amontillado by Edgar Allan Poe836 Words   |  3 Pagesqualities in the story. In the story many things are used as symbols such as the actual cask of amontillado, the trowel, the jester costume and the setting in which there is two in the story. Another literary technique used significantly in the story is irony. Irony is the expression of ones meaning by using language that normally signifies the opposite. In the short story â€Å"The Cask of Amontillado,† Montresor a very troubled man who plans to seek revenge on another man named FortunatoRead MoreEdgar Allan Poe And The Cask Of Amontillado1384 Words   |  6 PagesWhat makes Edgar Allan Poe work unique? Other than being a strange individual, Poe has become a remarkable literature writer. The Raven, Annabel Lee, and The Cask of Amontillado are just a few of Poe’s work that staples the theme of gothic literature. This essay will allow you to see the gothic elements Edgar Allan Poe uses through his most common poems. Gothic literature has many elements which play into its definition. The actual definition is a style of writing that is characterized by elementsRead MoreThe Cask Of Amontillado By Edgar Allan Poe906 Words   |  4 Pages â€Å"The Cask of Amontillado† â€Å"The Cask of Amontillado† was written in 1846, by Edgar Allan Poe. Born in 1809, Poe never knew any of his parents. At the age of three, his mother died of tuberculosis, and his father deserted the family before he was born. Taking care of him was his foster parents in Richmond, Virginia. They loved Poe, but were not supportive of his decisions and kept Poe poor. Having debt and not being able to provide food and clothes for himself caused Poe to quit school. Later, heRead MoreThe Cask Of Amontillado By Edgar Allan Poe920 Words   |  4 Pageswhen that trust no longer exists? In â€Å"The Cask of Amontillado† written by Edgar Allan Poe, Fortunato is about to find the answer to this question. On the surface, Montresor seems friendly with Fortunato, but deep down he feels nothing but hate for him. Could this hatred have an irrationality that only Montresor understands? In different ways, both of these men are proud and affluent, yet both have downfalls that will l ead to a tragic ending. Edgar Allan Poe’s use of language contributes to the understandingRead MoreThe Cask Of Amontillado By Edgar Allan Poe1555 Words   |  7 PagesIn his writing, Edgar Allan Poe has multiple uses of direct and indirect characterization. In The Cask of Amontillado, Montresor had rules such as â€Å"I must not only punish but punish with impunity. A wrong is unredressed when retribution overtakes its redresser. It is equally unredressed when the avenger fails to make himself felt as such to him who has done the wrong† (Poe, 2). Poe used indirect characterization to show the reader that Montresor is an unreliable narrator because he justified hisRead MoreThe Cask Of Amontillado By Edgar Allan Poe1303 Words   |  6 PagesIn Edgar Allan Poe’s â€Å"The Cask of Amontillado,† the narrator recalls an extremely significant time in his life, and takes the reader along with him. Throughout the story, one experiences a perfectly planned murder which took place over fifty years ago, and still no one has discovered what truly happened to poor Fortunato as he was chained to a wall in a room that was then closed off, and torched to death due to all the nitre in the walls. As the story goes on, the reader can see some of Poe’s unfortunateRead MoreThe Cask Of Amontillado By Edgar Allan Poe1076 Words   |  5 PagesThe short story, The Cask of Amontillado, written by Edgar Allan Poe is a story of terror and betrayal. Like many of Poe’s literary works, the story has a dark undertone with a theme of terror and depression. More than half a century ago, Marshall McLuhan argued that though Poe was fascinated by evil, the evil that he had in mind was not that of Calvinism, but that of the split man and the split civilization. In general, McLuhan was right, but in this instance Calvinism, and its God, provided a darkRead MoreThe Cask Of Amontillado By Edgar Allan Poe985 Words   |  4 PagesEdgar Allen Poe is a well kn own author of short stories and poetry from the 19th century. He is known especially for his stories of horror and suspense. The Cask of Amontillado is one of his more famous pieces. The story follows the narrator, Montresor, as he exacts revenge on Fortunato. Montressor draws Fortunato into the wine cellar where eventually he chains Fortunato to the wall and encloses him inside it. Throughout the story the narrator continually proves that he is not the most reliable sourceRead MoreThe Cask Of Amontillado By Edgar Allan Poe926 Words   |  4 PagesThe Cask of Amontillado, by Edgar Allan Poe, is a deliciously ironic story about guilty conscience. The central character, Montresor, has been insulted by his rival, Fortunato, and vows revenge. He plans to entomb Fortunato in the catacombs below his house, and he establishes a set of guidelines for a successful revenge. As the plan progresses, however, Montresor loses his determination. Though Montresor does entomb Fortunato, he cannot s avor the revenge because a guilty conscience consumes him

Truck Driving How to Do It Free Essays

How many of us can drive a truck? Most people wouldn’t know where to start. All they see is a huge chunk of metal with wheels and can’t fathom driving it. Using this guide will give a crash course (no pun intended) in truck driving and what it’s like on the road with these monsters. We will write a custom essay sample on Truck Driving: How to Do It or any similar topic only for you Order Now It also may help one understand the world of the truck driver and give them a new found respect for them. One of the first steps every truck driver must do before setting out on a trip is the pre-trip inspection. On some tractor-trailers there are as much as one hundred items that need to be checked or replaced. Some of these items include wheels and tires, brakes, lights, and fuel containers. All of which can be a major hazard to other drivers if they are not fixed or properly secured. Another equally important step for a truck driver is making sure the cargo is properly stowed or locked down. Depending on the trailer type there are various ways to properly secure the cargo, such as straps, chains, and wedge blocks. If the cargo isn’t properly secured it can shift and cause the truck to be overweight in a certain area. Being overweight on a certain side or on a certain axel can have a devastating effect on the road or on the truck itself. Federal dictates the no tractor-trailer can exceed 80,000 lbs without proper paperwork. Another area that needs to be checked thoroughly is the inside of the cab and the dash instruments. Most trucks have three times as many gauges as a pedestrian vehicle. Some that you would not find in a car is the air (psi), exhaust temp. , turbo pressure, and oil temperature. It is very important that we keep a check on all the various fluids the truck needs as well. Most trucks will not crank if a fluid level is to low such as the water. Once out on the road a truck driver has to be extremely cautious and aware of what is happening on the road. One of the most overlooked and easiest things to do is watching the mirrors very often. The mirrors are there to help you drive safely and efficiently. It is a very important step to master for the safety of ourselves and the other drivers. There are many more steps and regulations to go along with truck driving. Each state has its own rules and regulations as well as certain cities. Most of which can be viewed on the state DMV website. I hope this essay helps anyone looking to become a truck driver, and remember, keep an eye on those mirrors. How to cite Truck Driving: How to Do It, Essay examples

La Machine a Rajeunir free essay sample

As soon as the caravan was on the road, we came across a police car. Five seconds later, another one passed us, its siren blaring loudly. Jo and I were sitting at the back on a car seat. Hugo and Sophie were facing us. Luce, on another seat, was reading an information book. We finally escaped the police. But this was not the end of our problems, like Jo already said. The assault of a police officer was a crime, and our escape made the situation worse. A1 the officers in the region were surely looking for us. But what worried me most was the helicopter. Why were he bandits trying to take my parents? What were their intentions and why are they interested in the phenomenon of their rejuvenation? For sure, these mysterious people are well organized. I already knew that the black car was following my dad and mom before they left. We will write a custom essay sample on La Machine a Rajeunir or any similar topic specifically for you Do Not WasteYour Time HIRE WRITER Only 13.90 / page Now, I know for sure that the spying started during their trip to Perc ©, when they left the machine of rejuvenation. Even if it seems stupid, the name of the machine is more than to Just attract tourists. Hugo was right. The machine was really capable of making humans younger. I could see things more clearly Dad and mom fell in the trap of a monstrous organization. They used them like guinea pigs for a strange experiment. Everything makes sense! The people in the helicopter were trying to capture my parents to analyze them, for sure! They must get to Perc © very quickly. If the machine of rejuvenation is the cause to our problems, then there could be a solution! When we arrive at Perc ©, we must be very careful. The bandits will surely try to kidnap my parents again. With her head lying against her small husband, mom was sleeping. l, who believed that the rejuvenation would stop, was well aware that not only would it continue, the process would speed up.