Cooking with biomass fuel and cardiovascular disease: a cross-sectional study among rural villagers in Phitsanulok, Thailand

Background: Burning biomass fuel is a major source of indoor air pollution; about 40% of Thai people still use biomass for cooking. There is increasing evidence of the association between biomass smoke exposure and serious health effects including cardiovascular disease. The object of this cross-sectional study was to investigate the association between biomass use for household cooking and cardiovascular outcome, including coronary heart disease, hypertension, high cholesterol, diabetes mellitus, and stroke among rural villagers in Phitsanulok, Thailand. Methods: Data from 1078 households were collected using a face-to-face interview questionnaire. In each household, data on cardiovascular disease, cooking practices, and cooking fuel, types of fuel they normally used for cooking, were collected. Results: After being adjusted for gender, age, cigarette smoke, secondhand smoke, and exposure to other sources of air pollution, it was found that the family members of cooks using biomass fuel were at risk of coronary heart disease (CHD; OR=4.35; 95%CI 0.10–18.97), hypertension (OR=1.61; 95%CI 1.10–2.35), high cholesterol (HC; OR=2.74; 95%CI 1.66–4.53), and diabetes (OR=1.88; 95%CI 1.03–3.46). Compared to LPG use, using wood was associated with stroke (OR=7.64; 95%CI 1.18–49.61), and using charcoal was associated with HC (OR=1.52; 95%CI 1.04–2.24). Compared to never user, household cooks who sometimes use charcoal had an increased risk of hypertension (OR=2.04; 95%CI 1.32–3.15), HC (OR=2.61; 95%CI 1.63–4.18), and diabetes (OR=2.09; 95%CI 1.17–3.73); and cooks who often use charcoal had an elevated risk of stroke (OR=3.17; 95%CI 1.04–9.71), and HC (OR=1.52; 95%CI 1.02–2.27) to their family members. Conclusions: The study results were consistent with those found in studies from other parts of the world, and supports that exposure to biomass smoke increase cardiovascular diseases. The issue should receive more attention, and promotion of clean fuel use is a prominent action.


Introduction
Cooking smoke is a major source of household air pollution, which affects billions of people around the world, especially in developing countries. Globally, nearly 3 billion people still use solid fuels (wood, charcoal, crop residues, and dung) for cooking and heating 1 . Smoke from wood burning contains a large number of pollutants, including particulate matter, carbon monoxide, nitrogen dioxide, formaldehyde, and a number of highly toxic organic compounds, such as benzene, 1, 3 butadiene, benzo[a]pyrene and other toxic polycyclic aromatic hydrocarbons 2 . In addition to fuel burning smoke, overheated of cooking oils might also produce smoke which depended on several factors, including cooking oils, cooking methods, cooking equipment, and food types 3 .
The use of solid fuel for cooking and/or household energy sources increases respiratory and non-respiratory illnesses in both adults and children. Those effects that are well established are acute respiratory infections, chronic obstructive pulmonary disease (COPD), lung cancer, asthma, tuberculosis, and cataracts 4,5 . In children, biomass use is related to mortality, and acute lower respiratory tract infections, and some other non-respiratory illness, such as poor lung function, low birthweight, nutritionaldeficiency, and impairment of learning ability 6,7 .
Though with limited evidence, recent studies linked biomass smoke exposure and cardiovascular diseases (CVD), e.g. coronary heart disease (CHD), hypertension, diabetes, and stroke [8][9][10][11] . In laboratory studies, chronic exposure to biomass smoke increased the thickness and plaque of blood vessels 12 . In epidemiological studies, Peruvians who live in high altitude environments and use biomass fuel had an elevated prevalence of hypertension 13 . A study among villager women in Bangladesh reported an association between elevated cumulative exposure to biomass smoke and the prevalence of hypertension 14 . A similar result was found in a study in Shanghai Putuo, which found using solid fuel increases the risk of hypertension, CHD, and diabetes 15 ; and a study in Shanxi, China reported an increased risk of hypertension, CHD, stroke, diabetes, and dyslipidemia 16 . A recent study by Yu et al. 17 also linked solid fuel use to cardiovascular mortality.
On a global scale, CVD is the number one cause of death and is responsible for about 18 million deaths annually 18 . In Thailand, CVD accounts for 23% of the national mortality 19 . Currently, there is no study on the effect of biomass smoke on CVD in Thailand. It was reported that about 40% of Thai households still use biomass, mainly charcoal, wood, and agriculture residue, for cooking 20 . The objective of this study is to investigate a possible association between biomass use for cooking and cardiovascular diseases, including CHD, hypertension, HC, diabetes, and stroke. The study uses data from a cross-sectional survey among rural villagers in Phitsanulok, Thailand. The result could be used for disease prevention and control, and to support the global literature.

Study design and setting
This is cross-sectional study. Participants are rural villagers living in Phitsanulok Province, Thailand. Phitsanulok is a midsize province located about 400 km north of Bangkok. There are 866,891 people in the area of 9 districts. Most of the people are rice farmers 21 .
Study participants and sampling procedure Participants were randomly selected using multistage sampling. Out of the 9 districts in Phitsanulok province, 5 were randomly selected. In each district, one sub-district and a local health-promoting hospital were approached. In each sub-district with support from the local health-promoting hospital, a total of 1,150 households were approached and 1,134 (98.6%) people agreed to participate in the study. In each household, only one participant who was responsible for household cooking and aged over 20 years was selected. After data cleanup, 56 (4.9%) items of data were missing important information, such as age, gender, cooking practice. The final data from 1,078 people were used for statistical analysis.
The minimum sample size was calculated to be 1,034, using unmatched cross-sectional study with the following assumptions:two-sided significance level = 95%; power of detection = 80%; percent unexposed with outcome = 5%; and odds ratio = 2.0.

Study questionnaire
Data was collected using a face-to-face interview questionnaire, which was administered by 15 village health volunteers (provided as Extended data in English 22 ). The interviews took place in the house of participants. The data was collected during the period of May-June 2017. Health volunteers were all trained on how to properly carry out the interview and use the questionnaire. The questionnaire was designed to collect information on demographic data, fuel use for cooking, and other cooking practices. In addition to general demographic data, participants were also asked a history of tobacco use (ever, never), and working in factory environments using "yes" or "no" questions. Ever smoker referred to those who smoke more than 100 cigarettes in their lifetime. Data on pesticide use was also measured by "yes" or "no" questions: "Have you ever spray or mix pesticide?". For cooking fuel data, we asked about the types of fuel they used for cooking food (wood, charcoal, LPG, electricity), and the frequency of using each types of fuel. Data collected on cooking practices were types of cooking oil, the frequency of tears while cooking (TWC) (never, sometimes, often), kitchen location (inside a house, outside a house, both inside and outside a house), and the characteristics of kitchen ventilation (good or poor ventilation).
The presence of cardiovascular disease was determined by the participant response to the question: "Have you ever been diagnosed with the following diseases (coronary heart disease (CHD), hypertension, high cholesterol (HC), diabetes mellitus, stroke) by a medical doctor?". For diseases among their family members, we asked "Did you have a family member with the following diseases?".
The content validity of the questions was tested by three experts, and the Index of Item Objective Congruence (IOC) was between 0.7-1.0. The questionnaire was also tested for question sequencing and understanding using a group of 30 people with a similar background to the intended participants.

Statistical analysis
Demographic and prevalence of cardiovascular disease were descriptively analyzed. Comparison between groups were analyzed using chi-square test for categorical variables, and independent t-test for continuous variables. The association between cardiovascular disease was analyzed using binary multiple logistic regression with odds ratios (OR) and 95 percent confidence interval (CI) adjusted for gender (male, female), age (continuous data), cigarette smoking (ever, never), living smoker (yes, no), working with smokers (yes, no), and exposure to air pollution (yes, no). These adjusted variables of the repondents were used also when analysis for ORs of disease risk among the respondents' family members. All statistical analyses were performed using IBM SPSS version 19 and OpenEpi (online version 3.01). Statistical significance was set at a p-value of less than 0.05.

Ethical considerations
The study was approved by the Ethical Committee of Naresuan University (COA No. 485/2016), and written informed consent from the respondents was obtained before the interviews were conducted.

Results
Most of the respondents were women (84.2%) with a mean age of 53.04 ± 12.93 yr. The highest education levels were primary school or high school. Most were farmers (36.0%) and 20.2% were causal workers on farms. About 10% were smokers and 33% lived with a smoker. Additional information on the demographic data is shown in Table 1 and in Underlying data 23 .
About 70% of the respondents reported using biomass for cooking (Table 2). However, when asked for fuel types that they usually use for cooking, 64.5% reported LPG and 32.3% charcoal. Among those who use charcoal, 38.6% use it often. About half have a kitchen located inside a house with good ventilation. Almost all reported having TWC either sometimes or often. Most of them cook every day.
The study found hypertension, HC, and diabetes to be the most common cardiovascular outcomes (Table 3). Compared to non-user group, biomass users had a significantly higher prevalence of hypertension, and HC, and their family members also had more incidence of hypertension, HC, diabetes, and heart disease. Further analysis using logistic regression and control variables, revealed that compared to gas users, biomass users had family members with elevated CHD, hypertension, HC, and diabetes (Table 4). Among different types of fuel, household cooks using wood had a significant elevated risk of CHD (OR=7.64, 95%CI 1.18-49.61), and their family members had an elevated risk of HC (OR=1.52, 95%CI 1.04-2.24). Comparing frequency of charcoal use, those who use charcoal sometimes or often are more likely to have CHD, hypertension, HC, and diabetes as compared to those who never use charcoal. The family members of charcoal users also had a significant increase of HC and stroke. When using TWC as an indicator for smoke exposure, it was found that those who always had TWC had significantly increased risk of stroke (OR=2.16; 95%CI 1.08-4.32), and those with sometimes TWC had a CHD risk (OR=2.64; 95%CI 1.02-6.81). Regarding kitchen location, the family members of cooks having kitchens both inside and outside a house had an elevated risk of stroke (OR=4.60; 95%CI 1.14-18.54).

Discussion
This study presented an association between cardiovascular diseases and exposure to smoke from biomass, mainly charcoal, which is relatively cleaner when compared to wood, coal, or dung, a biomass which were often found in the literature. The study also showed that biomass use not only affects household cooks but also their family members. It was found that biomass users have a higher prevalence of hypertension and HC, and their family members had a higher prevalence of hypertension, HC, diabetes, and CHD (Table 3). Further analysis using logistic regression and control for potential confounder showed a significant OR of biomass use and CHD(F), hypertension(F), HC(F), and diabetes(F) ( Table 4). Compared to LPG, wood use also had a strong association with stroke (OR=7.64; 95%CI 1.18-49.61). Among charcoal users, those who use it sometimes or often had an elevated risk of CHD, hypertension, HC, and diabetes for themselves, and risk of HC and stroke for their family members. The results are consistent with the literature. Previous research found biomass smoke contains a lot of pollutants, especially fine particulates, and carbon monoxide which are known to cause cardiovascular effects 2 . In laboratory studies, biomass smoke exposure was associated with endothelial inflammation 24 .
For hypertension, we found both cooks and their family members have a higher prevalence of hypertension (Table 3).     The current study also found a higher prevalence of HC among cooks and their family members using biomass fuel ( We found about 10% of the respondents had type 2 diabetes and the prevalence of the disease was higher among biomass users ( Those who use biomass for cooking had a risk of CHD 4.35 times (95%CI 0.10-18.97) of LPG users; and those using charcoal sometimes had risk of CHD 4.11 times (95%CI 1.40-12.11) of never user group. These results are consistent with evidence from cigarette smoke and ambient air pollution.
In animal studies, biomass fuel smoke caused arteriosclerotic effects in animal blood vessels 12 . Studies found COPD as a risk factor of CHD 31 ; and our previous study found elevated chronic symptoms, such as chronic cough, dyspnea and runny nose which is a sign of COPD among cooks using biomass fuel for cooking 3 . Epidemiological studies also reported an association between solid fuel smoke exposure and CHD 32 . A study in Pakistan found that rural women who currently use solid fuel had an increased risk of acute coronary syndrome (OR=4.8; 95%CI 1.5-14.8) 33 . This is consistent with a study from Shanghai Putuo, which found solid fuel use in the home is associated with CHD (OR=2. One potential drawback of this study was the use of self-reported data of diseases. Without the confirmation of medical records, the survey diseases are subjected to information bias. However, the bias will be distributed equally to all comparison groups, and this tends to underestimate the result. The number of participants included in this study was also rather small to detect the actual association of a rare disease, e.g. stroke. By using cross-sectional design, the study result cannot explain the causal relationship, because it is not known whether exposure or the disease occurred first. However, the problem is minimal for rare diseases.

Conclusions
The results from this study support research findings in other part of the world that using biomass for cooking increases the risk of cardiovascular diseases. This study also confirms the negative effects of using charcoal, which is considered to be a relatively cleaner fuel as compared with wood, dung, coal, and other agricultural residues. Concerned organizations should pay more attention to the issue and promote clean fuel usage. First off, thank you for setting out to provide more evidence on this important public health issue that affects a large proportion of people globally. I will categorize my comments under major and minor issues with the manuscripts that I believe should be worked on.

Major issues:
I believe there is a need for reanalysis of the data based on the issue I have raised in the result section concerning the definition of the biomass use which is a major variable of interest for this study i.e. classification of participants into biomass users and non-biomass users. This may probably result in a significant change in the results outputs, discussion and conclusion of your study.

Minor issues:
I am going to make my feedback based on the structure of the manuscript. Some of my comments have been made by the other peer reviewers but seems to have been missed in this most current version.

Abstract:
In the abstract, it may be good to write in full LPG. I believe it is best to use standardized abbreviations. I am not sure that HC is a widely used abbreviation?
○ "The object of this cross-sectional study was ……." ○ Do you mean "The objective of this cross-sectional study was ……."?
"In each household, data on cardiovascular disease, cooking practices, and cooking fuel, types of ○ fuel they normally used for cooking, were collected." Can be better phrased as: "In each household, data on cardiovascular disease, cooking practices, cooking fuel and types of fuel they normally used for cooking were collected." "…and supports that exposure to biomass smoke increase cardiovascular diseases." ○ Do you mean "…and supports that exposure to biomass smoke increases the risk of cardiovascular diseases."?

Introduction:
"In addition to fuel burning smoke, overheated of cooking oils might also produce smoke which depended on several factors, including cooking oils, cooking methods, cooking equipment, and food types3" ○ This sentence needs to be rephrased to ensure better clarity.
"non-respiratory illness, such as poor lung function, low birthweight,nutritionaldeficiency, and impairment of learning ability6,7" Space the bolded word as: "non-respiratory illness, such as poor lung function, low birthweight, nutritional deficiency, and impairment of learning ability6,7". ○ "….still use biomass, mainly charcoal, wood, and agriculture residue, for cooking" Can better read as: "….still use biomass mainly charcoal, wood and agriculture residue for cooking". "This is cross-sectional study." Can better read as: "This is a cross-sectional study". ○ I observed that the questionnaire was translated. What was the process used in the translation and maintaining its validity? I believe you need to address this.
"Ever smoker referred to those who smoke more than 100 cigarettes in their lifetime." Can better read as: "Ever smoker referred to those who have smoked more than 100 cigarettes in their lifetime." "About 70% of the respondents reported using biomass for cooking (  Again the data is found in Table 1 and not Table 2, except you rename all your tables.  Be consistent in using one term for people not using biomass as in some places you used non-user group and in some others, gas users. Meanwhile did you aggregate electricity users and LPG users? If so you may need to state it. ○ "..and their family members also had more incidence of hypertension, HC, diabetes, and heart disease." ○ Please replace as this study cannot assess for incidence but for prevalence. "..and their family members also had more prevalence of hypertension, HC, diabetes, and heart disease." ○ In Table 4 put a superscript at R and F to explain what it is like in Table 3. It would have been good to label the continuation of Table 4.
○ Is there any reason why you did not control for occupation and education in the logistic regression? Or don't you think they potentially can influence the results?
○ Discussion: "This study presented an association between cardiovascular diseases and exposure to smoke from biomass, mainly charcoal, which is relatively cleaner when compared to wood, coal, or dung, a biomass which were often found in the literature." If you planned to concentrate on charcoal then you should have clearly stated it and brought it out even in the result sections. It appears you may not have adequately brought this out in the write up and result so far. Again, any reference to show the statement you made that charcoal is cleaner than the others? I believe that is a statement that will need to be backed up with evidence and also should have been in the introduction.

Further analysis using logistic regression and control for potential confounder showed a significant OR of biomass use and CHD(F), hypertension(F), HC(F), and diabetes(F)
I think it may be good to write the bolded in full. Chudchawal Juntarawijit, Naresuan University, Phitsanulok, Thailand

Response to reviewer
In the abstract: Comments: The methods should be improved, giving a clear description of cooking fuel.

Responses:
A short description of cooking fuel was added to the methods section. Because F1000Research has set a maximum limit of 300 words for abstract, no more detailed information could be added.
Comments: The author mentioned 'hypertension' in the objective, but high blood pressure (HBP) in the result section. Terminology should be consistent throughout the manuscript.

Responses:
The term "high blood pressure" was replaced by "hypertension".
Comments: In the conclusion, the author stated, 'the study results were consistent with those found in studies from other parts of the world...'. This statement should be discussed in the discussion, not in the conclusion. The conclusion should be based on the authors' main findings.
Responses: Yes, I agree that "the statement should be discussed in the discussion". However, we believed we had already done that enough to justify the statement, which is our main finding.

In the introduction:
Comments: Please clarify the term 'Cooking smoke,' whether it means smoke from cooking or from fuel?

Responses:
The meaning of cooking smoke was clarified and more information was added to the first paragraph in Introduction.

In the methods:
Comments: The study design is well articulated. But self-reported cardiovascular diseases may give a vague impression to the readers, although the author mentioned it as a limitation. In a matter of sense, the authors showed more than two-thirds (71.9 %) of participants had primary education who acted as self reporters of disease condition. So, in my opinion, the authors should have a strong justification in favor of including self-reported cardiovascular diseases with authentic scientific references.
Responses: Yes, I agree that using self-report data is a limitation of this study. However, since the data was collected by a well trained and experienced village health volunteer, the problem was expected to be minimal. The quality of the answer to this question may not depend much on their background education of respondents. In addition, this information bias, if occurred, will equally distribute among groups (case and control).
Comments: Again, do the authors cross-check self-reported disease conditions with the patient's medical or laboratory reports or drug history? The mentioned high cholesterol (HC) is instead a biochemical abnormality apart from a disease condition.

Responses:
Yes, it is good if we can do the cross-check self-reported conditions. However, we did not do that.

In statistical analysis:
Comments: The regression model needs a precise description. Is it a multivariate or multinominal model?
Responses: Thank you for reminding.
In this study, we use binary multiple logistic regression.
More detail of the model was added to the statistic description.
Comments: In a logistic regression model, the cardiovascular disease condition of family members are also encountered. Are the adjusting confounding variables like age and sex in that regression model in relation of family members' age and sex, or the respondents'? It should be precisely mentioned in description of regression model.
We confirm that we have read this submission and believe that we have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.
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