MÉTODO BINOMIAL DE VALORACIÓN DE OPCIONES

Ethical Approval

Ethical approval for the project was obtained from ethical committee at Epsom NHS Trust (Appendix 1).

Subjects and Recruitment

A cohort o f 2300 children bom between 1 April 1995 and 31 April 1996 (13 months) at Epsom General Hospital in the Mid - Surrey area comprised the preliminary sample. Their details were available from Epsom NHS Trust Information Department. This included name, surname, address and date o f birth. The parent(s) were contacted when their children were 6 - 1 2 weeks old and invited to include their children in the study. This was done by personal contact at Child Health Clinics or by mail. In order to avoid contacting parents, whose children might have died after birth, the list o f the names was regularly updated through the List Cleaning Services at the Office o f National Statistics. Registration with the Office of National Statistics was required for the reasons of confidentiality (Appendix 2). An advertisement poster was designed and distributed in the area to invite families whose details might not have been available, for example those bom at home or out o f the area (Appendix 3). The Health Visitors in the area were informed o f the study.

An information sheet and consent form were handed or sent to the parent(s) / carers of children (Appendix 4 and 5). The parent(s) / carers were asked to sign the consent form if they were willing for their children to participate in the study. A reminder letter was sent to non- respondents. It was not possible to follow up by telephone, as the family telephone number was not available.

The parents participating in the study were required to carry out the following tasks: (1) Completion o f a prepared 3 - day food diary for their children at 6, 12, and 18

months of age.

(2) Attendance for a dental examination at 12 and 18 months o f age. (3) Recording of the time of deciduous incisor teeth eruption (Appendix 6)

The parents, their health visitors, and dental staff at Epsom NHS Trust were kept informed on the progress o f the study, by annual Newsletter.(Appendix 7). To thank parents for their help and support a specially designed greeting card was sent to them at Christmas 1997.

Description of the Area

Mid - Surrey area where the study was carried out is located within 30 miles of Central London to the South West with a population o f approximately 167,400 according to 1991 Census (O.P.C.S. 1991). The 1991 Census revealed that 68.6 per cent o f men aged 1 6 - 6 4 and 47.2 per cent o f woman aged 1 6 - 5 9 resident in the M id- Surrey were in employment (Khakoo 1993). Five per cent o f men and four per cent of women were unemployed. Overall, people living in the area are fairly affluent with over fifty per cent o f employed Mid - Surrey residents belonged to Social Classes I and n at 1991 Census (Khakoo, 1993). At 1991 Census, 96 % of the population were White (Khakoo 1993).

The birth number in 1991 was 1902. The infant mortality rate for children under 1 year of age was 7.3 per 1000 live birth (Khakoo 1993). The projected birth number for

1995 was 2600. Epsom NHS Health Care Trust is the main health service provider in the area. Within the Trust there are 9 clinics from which Health Visitors care for infants resident in their district: Banstead, Epsom, Ewell, Leatherhead, Tattenham, Cobham, Bourn Hall, Ashtead and Great Bookham. These are located within 2 - 1 5 miles of each other. There are five Dental Surgery Clinics within the Trust at Banstead, Epsom, Ewell, Leatherhead, and Tattenham Health Centres. The water in Mid - Surrey area is not fluoridated and the fluoride level of water ranges between 0.1 to 0.2 ppm in different localities.

The information on dental health of pre -school children in the area is lacking. Nevertheless, the information on dental health o f 5- year- olds carried out by NHS program o f caries prevalence studies co -ordinated by the British Association for the Study o f Community Dentistry (BASCD) between September 1997 and April 1998 was available (Harris 1998; Pitt and Evans 1998). Thirty two percent o f 5-year-old children

decay, and the mean decayed teeth (dt) for this group o f children was 3.4 (Sd 2.6) teeth. Whereas, the mean (dt) for all children was 0.92 (Sd 2.0).

Materials

1 - Tooth Eruption Form

A tooth eruption form was designed for parents to keep a record o f the date as each incisor tooth erupted (Appendix 6). It was sent with a pre - paid envelope to parent(s) after they agreed to include their children in the study. The parent(s) returned the form after all incisors teeth erupted and the dates were recorded.

2 - Longitudinal Diet Diaries

A 3 - day food diary was designed based on the diet sheet used in a previous study (Roberts et al. 1993). The diary was further revised and piloted on a group of parents with children o f similar age to the study group (Appendix 8). The diaries were sent to parent(s) / carer with a pre-paid envelope at approximately 2 weeks before their children were 6, 12 and 18 months of age. Parent(s) / carers were asked to record food intakes using household measures. They were asked to choose 3 days (2 weekdays and 1 weekend day) when their children were eating normally and begin recording after midnight by writing down the time at which any food and / or drink were taken. They were also zisked to record how long breast or bottle-feeding had taken and also to either weigh the food the child had taken or describe them in handy measures (Appendix 8). The instructions were included in the diary. The non-complaints were followed up.

3 - Questionnaire

A questionnaire was designed to provide detailed longitudinal information on feeding practice, oral hygiene behaviour and demographic background o f children (Appendix 9). It was first piloted on a group o f mothers with children at approximately similar age to the study group, and then was further revised and used in the study. The questionnaire had three areas of emphasis:

Detailedfeeding behaviour from birth: questioning the feeding method, frequency and pattern of feeding, type, age and duration for using different milks, bed time and night time feeding, type o f bottle and drink vessel used during day time and night time, use of

Oral hygiene behaviour, questioning age tooth brushing started, frequency, time of brushing, whether toothpaste used or not, type o f toothpaste, and whether fluoride supplements used, and duration

The sociodemographic background: questioning parents age, education, race, ethnic origin, occupation, whether parents received any social benefit, and whether both parents were working, the child care arrangement when they were at work as well as the child’s gender and birth order.

After each dental examination parents / carers were asked to complete the questionnaire and return it by post.

Dental Exam ination

The families who participated in the study were invited to have a dental examination carried out on their children at 12 and 18 months of age. Dental examinations

continued for 18 months between April 1996 and November 1997 when the participants were 12 and 18 months old. The dental examinations were carried out at Health Centre Clinics in the Epsom NHS Trust. Two reminders were sent to non - attendants.

Dental Exam ination Procedure

A dental appointment was arranged at one o f the Health Centres in the Trust, where the parents usually attended for their children’s routine health surveillance. The

appointment was sent to the parent(s) when each child was due for dental examinations at 12 and 18 months o f age.

When the dental examination was carried out at dental surgery clinics the parent sat on the dental chair and the child lay on the parent’s lap. At clinics with no dental chair the child was examined on lap position by lying on examiner’s lap but facing toward the parent(s). The examiner (MH), who was calibrated against an experienced community dental officer, examined all children. Only a mouth mirror and light were used. If the standard dental light was not available a hand held torch [ Daray lighting ltd, 7

CommerceWay, Stanbridge Road, Leighton Buzzard, Beds ] was used. The dental examination covered the followings:

present in the mouth. A plaque sample was removed from the buccal surface o f the upper central incisors by using a sterile toothpick. The toothpick was moved against the tooth surface gingivo-incisally. The plaque samples were immediately placed into 1 ml o f Fastidious Anaerobe Broth (FAB) supplemented with 30 percent glycerol and

transported to the laboratory. They were stored at -70° C until required for bacteriological examination.

Bacterial Dental Plaque

Each quadrisection (mesiobuccal, distobuccal, distolingual, mesiolingual) o f all erupted teeth surfaces were examined for the presence o f bacterial dental plaque. A simple index was used to estimate the number of tooth surfaces which had discernible deposits of bacterial dental plaque (O’Leary, 1972). A simple presence or absence o f plaque was used. It was not possible to use more detailed plaque and gingival indices. Very young children are unable to co- operate for the period o f time necessary for more detailed examination. The results were recorded on a specially designed form.

Gingivitis

The gingivae were examined visually for inflammation, determined by the presence or absence of reddened gingival tissue. The areas of gingiva recorded were the same as for plaque (mb, db, dl, and ml) on each quadrisection o f the erupted teeth (O’Leary 1972, Franco et al. 1996). The results were recorded on a specially designed form

Dental Caries

The teeth were cleaned and dried Nvith a cotton wool roll and all surfaces (mesial, buccal, distal, lingual, and occlusal) were visually examined for dental caries. A mouth mirror and light was used as diagnostics aids. The World Health Organisation criteria (WHO 1987) were used for the diagnosis o f caries.

Microbiological Methods

The samples were shaken with sterile glass beads, and diluted in Fastidious Anaerobe Broth (FAB, LabM, Bury, Lancs, UK) by transferring a 100 pi aliquot o f sample to 900 pi of FAB to make a 10'* dilution. Then 1 in 10 again to produce a 10 dilution neat of the sample. A 100 pi aliquot was spread onto BMSA [ Mitis - Salivarius agar (Difco)

(Oxoid, Basingstoke, Hants,Uk) and Sabouraud Dextrose agar (Oxoid), using a sterile hockey - stick shaped spreader. These media were used for the enumeration o f mutans streptococcus, lactobacilli and yeasts, respectively. A small number o f the samples were also plated onto Fastidious Anaerobe Agar supplemented with 5 % (V/V) horse blood at dilutions of neat: 10 and 10“^ to determine the viability o f the bacteria following storage.

The BMSA and Rogosa media were cultured in anaerobic chamber in an atmosphere of N]: 80°: CO2 : H2 : 10 for 3 days, and the Sabouraud Dextrose Agar was incubated for 3 days aerobically. The Sabouraud Agar is a selective media for yeasts, which appear as creamy smooth or slightly wrinkled colonies. The number of mutans streptococci was recorded from the BMSA media, on which they form characteristic dark blue colonies, which appear firmly adhered to the Agar. Polysaccharide may be observed as a drop of liquid on top o f the colony. Lactobacilli were enumerated from the Rogosa Agar on which they appear as large to small cream colonies. On the basis o f these counts the numbers of each taxa were calculated in the original sample and were expressed as colony forming units (cfu) per plaque sample.

Calibration and Reproducibility

Calibration studies for diagnosis o f dental caries and detection of visible plaque were carried out in a Playgroup and included 25 children aged between 2 . 5 - 3 years, by re­ examining them within 1 week intervals.

Data Processing and Analysis

All dental, dietary and bacteriological data were coded and entered onto a especially designed database on Access Microsoft.

Dietary Data

Each returned food diary was checked for completeness by a research nutritionist and descriptive terms were converted into weight (Food Portion MAFF 1993).

manufacturers excluded total sugar composition, therefore types o f sugar in the di:t could not be quantified. The total carbohydrate intaikes were produced by using Food Table and the information from baby food manufactturer companies, when available. Some manufacturers (particularly some baby food nnanufacture) did not provide this information on the amounts of sugar and starch conttents. Therefore, the calculated percentage o f energy intake from sugars and starch (did not add up to the percentage of energy intake provided by the carbohydrate.

One observer calculated frequencies o f non-milk extrinsic sugar episodes per day. Non-milk extrinsic sugars are all sugars not containied within the cellular structure of the food, whether natural or refined, excluding lactose in milk and milk products (Department o f Health, 1989). Formula milk and soya milk containing glucose / maltodextrins were included into this category. An episode was defined by an occasion when drink, food or both were consumed within one of the allocated time periods, or at least a 30 minute time period had elapsed between tlhe previous consumption o f food / drink. Total eating / drinking occasions per day w ere also calculated using this method and categorised into 18 various food / drink groups (described in Appendix 11. All data were then entered onto a database on Access Microsoft and were then analysed using SPSS™ for Windows Release 6 (Norussis 1996) a c(omprehensive statistical analysis package.

Dental Data

All dental data were entered onto a database on Aiccess Microsoft. The number o f teeth quadrisections covered with plaque was talliedl as (plaque score). The proportion o f surfaces covered by plaque as the percentage o f alll surfaces present was calculated as (plaque index). The same calculation was used for gingival score and index. Children with plaque / gingival score zero were grouped as (plaque free / gingivitis free) and those with plaque / gingival score more than zero as (with plaque / with gingivitis)

Bacteriological Data

Children from whom mutans streptococci, lactobacilli or yeasts were not isolated, were recorded as negative. For these taxa a value off 0 was used in subsequent analysis. The microbial counts were transformed to log lo (cfu 4-1).

Statistical Analysis of All Data

All data were analysed by using SPSS. All raw and loglo transformed data were tested for normality using the appropriate tests (Shapiro - Wilks test). The majority of data were not normally distributed, therefore non - parametric tests such as Mann - Whitney U test or Kruskaul -W allis tests were used for comparison between groups. Spearman rho for correlation between variables was carried out, too. For normally distributed data the appropriate parametric test such as t - test, and Pearson test were under taken. Chi square test was used for the comparison o f different variables between groups. Statistical level o f significant was set at p < 0.5.

CH A PTER 4

RECRUITM ENT AND SUBSAMPLE SELECTION

The recruitment continued for 14 months between April 1995 and June 1996. The children were between 4 and 12 weeks old at recruitment. Table 4.1 shows that a total o f 1380 (60 %) parents who were invited, consented for their children to participate in the study. A total o f 71 (3 %) parents declined to let their children take part in the study and 849 (37 %) did not reply (Table 4.1).

Drop outs

The study lasted for 2 years and 6 months between the time when the recruitment started and the time when the last dental examination was completed. During this period, o f 1380 children whose parents consented, a total o f 191 failed to complete the study. This was due to 120 (9 %) who moved away from the area and 69 (5 %) who dropped out and 2 children who died (Table 4.2). Therefore, a total o f 1189 children comprised the sample when the study came to an end after the completion of the final dental examination (Table 4.2). Nevertheless, not all parents and children co - operated with all parts o f the study. The response rate for each part is described.

Table 4.1 Response rates to recruitm ent

G roup Num ber O f C hildren Percentage

Parents invited 2300 100

Refusals 71 3

Non - contacts 849 37

Consent to the study 1380 60

Table 4.2 The num ber and percentage of children who moved out of the area or dropped out of the study

N um ber of children As % of all invited

1380 60%

Consent to the study

Moved away / dropped out Moved Dropped Total at: < 6 months o f age 29 11 40 6 - 1 2 months o f age 18 12 30 1 3 - 1 8 months of age 73 46 119 120 69 189 Death 2 Total 191 Participants 1189 52%