Ruminant-associated Listeria monocytogenes isolates belong preferentially to dairy-associated hypervirulent clones: a longitudinal study in 19 farms.


Journal

Environmental microbiology
ISSN: 1462-2920
Titre abrégé: Environ Microbiol
Pays: England
ID NLM: 100883692

Informations de publication

Date de publication:
12 2021
Historique:
received: 03 08 2021
accepted: 24 11 2021
pubmed: 5 12 2021
medline: 18 3 2022
entrez: 4 12 2021
Statut: ppublish

Résumé

Studies have shown that ruminants constitute reservoirs of Listeria monocytogenes, but little is known about the epidemiology and genetic diversity of this pathogen within farms. Here we conducted a large-scale longitudinal study to monitor Listeria spp. in 19 dairy farms during three consecutive seasons (N = 3251 samples). L. innocua was the most prevalent species, followed by L. monocytogenes. Listeria monocytogenes was detected in 52.6% of farms and more frequently in cattle (4.1%) and sheep (4.5%) than in goat farms (0.2%). Lineage I accounted for 69% of L. monocytogenes isolates. Among animal samples, the most prevalent sublineages (SL) and clonal complexes (CC) were SL1/CC1, SL219/CC4, SL26/CC26 and SL87/CC87, whereas SL666/CC666 was most prevalent in environmental samples. Sixty-one different L. monocytogenes cgMLST types were found, 28% common to different animals and/or surfaces within the same farm and 21% previously reported elsewhere in the context of food and human surveillance. Listeria monocytogenes prevalence was not affected by farm hygiene but by season: higher prevalence was observed during winter in cattle, and during winter and spring in sheep farms. Cows in their second lactation had a higher probability of L. monocytogenes faecal shedding. This study highlights dairy farms as a reservoir for hypervirulent L. monocytogenes.

Identifiants

pubmed: 34863016
doi: 10.1111/1462-2920.15860
doi:

Types de publication

Journal Article Research Support, Non-U.S. Gov't

Langues

eng

Sous-ensembles de citation

IM

Pagination

7617-7631

Informations de copyright

© 2021 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.

Références

Addis, M.F., Cubeddu, T., Pilicchi, Y., Rocca, S., and Piccinini, R. (2019) Chronic intramammary infection by Listeria monocytogenes in a clinically healthy goat - a case report. BMC Vet Res 15: 229.
Borucki, M.K., Gay, C.C., Reynolds, J., McElwain, K.L., Kim, S.H., Call, D.R., and Knowles, D.P. (2005) Genetic diversity of Listeria monocytogenes strains from a high-prevalence dairy farm. Appl Environ Microbiol 71: 5893-5899.
Carlin, C.R., Liao, J., Weller, D., Guo, X., Orsi, R., and Wiedmann, M. (2021) Listeria cossartiae sp. nov., Listeria immobilis sp. nov., Listeria portnoyi sp. nov. and Listeria rustica sp. nov., isolated from agricultural water and natural environments. Int J Syst Evol Microbiol 71: 004795.
Castro, H., Jaakkonen, A., Hakkinen, M., Korkeala, H., and Lindström, M. (2018) Occurrence, persistence, and contamination routes of Listeria monocytogenes genotypes on three Finnish dairy cattle farms: a longitudinal study. Appl Environ Microbiol 84: 1-14.
Cavicchioli, R., Ripple, W.J., Timmis, K.N., Azam, F., Bakken, L.R., Baylis, M., et al. (2019) Scientists' warning to humanity: microorganisms and climate change. Nat Rev Microbiol 17: 569-586.
Charlier, C., Perrodeau, É., Leclercq, A., Cazenave, B., Pilmis, B., Henry, B., et al. (2017) Clinical features and prognostic factors of listeriosis: the MONALISA national prospective cohort study. Lancet Infect Dis 17: 510-519.
Chow, J.T.H., Gall, A.R., Johnson, A.K., and Huynh, T.N. (2021) Characterization of Listeria monocytogenes isolates from lactating dairy cows in a Wisconsin farm: antibiotic resistance, mammalian cell infection, and effects on the fecal microbiota. J Dairy Sci 104: 4561-4574.
Clinical and Laboratory Standards Institute (CLSI) (2017) Performance standards for antimicrobial susceptibility testing - approved guideline M100, 27th ed.
Cooper, J., and Walker, R.D. (1998) Listeriosis. Vet Clin North Am Food Anim Pract 14: 113-125.
Costard, S., Espejo, L., Groenendaal, H., and Zagmutt, F.J. (2017) Outbreak-related disease burden associated with consumption of unpasteurized cow's milk and cheese, United States, 2009-2014. Emerg Infect Dis 23: 957-964.
Dell'Armelina Rocha, P.R., Lomonaco, S., Bottero, M.T., Dalmasso, A., Dondo, A., Grattarola, C., et al. (2013) Ruminant rhombencephalitis-associated Listeria monocytogenes strains constitute a genetically homogeneous group related to human outbreak strains. Appl Environ Microbiol 79: 3059-3066.
Doumith, M., Buchrieser, C., Glaser, P., Jacquet, C., and Martin, P. (2004) Differentiation of the major Listeria monocytogenes Serovars by multiplex PCR. J Clin Microbiol 42: 3819-3822.
Dreyer, M., Aguilar-Bultet, L., Rupp, S., Guldimann, C., Stephan, R., Schock, A., et al. (2016) Listeria monocytogenes sequence type 1 is predominant in ruminant rhombencephalitis. Sci Rep 6: 36419.
Esteban, J.I., Oporto, B., Aduriz, G., Juste, R.A., and Hurtado, A. (2009) Faecal shedding and strain diversity of Listeria monocytogenes in healthy ruminants and swine in northern Spain. BMC Vet Res 5: 1-10.
EUCAST. (2020) Disk diffusion method for antimicrobial susceptibility testing, version 9.
European Union. (2010) Directive 2010/63/EU of the European Parliament and of the Council of 22 September 2010 on the protection of animals used for scientific purposes. Off J Eur Union: 33-79.
Fox, E., Hunt, K., O'Brien, M., and Jordan, K. (2011) Listeria monocytogenes in Irish Farmhouse cheese processing environments. Int J Food Microbiol 145: S39-S45.
Gray, M.J., Zadoks, R.N., Fortes, E.D., Dogan, B., Cai, S., Chen, Y., et al. (2004) Listeria monocytogenes isolates from foods and humans form distinct but overlapping populations. Appl Environ Microbiol 70: 5833-5841.
Grif, K., Patscheider, G., Dierich, M.P., and Allerberger, F. (2003) Incidence of fecal carriage of Listeria monocytogenes in three healthy volunteers: a one-year prospective stool survey. Eur J Clin Microbiol Infect Dis 22: 16-20.
Hafner, L., Pichon, M., Burucoa, C., Nusser, S.H., Moura, A., Garcia-Garcera, M., and Lecuit, M. (2021) Listeria monocytogenes faecal carriage is common and depends on the gut microbiota. Nat Commun 12: 6826.
Haley, B.J., Sonnier, J., Schukken, Y.H., Karns, J.S., and Van Kessel, J.A.S. (2015) Diversity of Listeria monocytogenes within a U.S. Dairy Herd, 2004-2010. Foodborne Pathog Dis 12: 844-850.
Ho, A.J., Ivanek, R., Gröhn, Y.T., Nightingale, K.K., and Wiedmann, M. (2007) Listeria monocytogenes fecal shedding in dairy cattle shows high levels of day-to-day variation and includes outbreaks and sporadic cases of shedding of specific L. monocytogenes subtypes. Prev Vet Med 80: 287-305.
Hurley, D., Luque-Sastre, L., Craig, T.P., Huynh, S., Eshwar, A.K., Nguyen, S.V., et al. (2019) Whole-genome sequencing-based characterization of 100 Listeria monocytogenes isolates collected from food processing environments over a four-year period. mSphere 4: e00252-19.
Hurtado, A., Ocejo, M., and Oporto, B. (2017) Salmonella spp. and Listeria monocytogenes shedding in domestic ruminants and characterization of potentially pathogenic strains. Vet Microbiol 210: 71-76.
Husu, J.R. (1990) Epidemiological studies on the occurrence of Listeria monocytogenes in the feces of dairy cattle. J Vet Med Ser B 37: 276-282.
Husu, J.R., Seppänen, J.T., Sivelä, S.K., and Rauramaa, A.L. (1990) Contamination of raw Milk by Listeria monocytogenes on dairy farms. J Vet Med Ser B 37: 268-275.
Hutcheson, K. (1970) A test for comparing diversities based on the shannon formula. J Theor Biol 29: 151-154.
Ianevski, A., Zusinaite, E., Shtaida, N., Kallio-Kokko, H., Valkonen, M., Kantele, A., et al. (2019) Low temperature and low UV indexes correlated with peaks of influenza virus activity in northern Europe during 2010-2018. Viruses 11: 207.
Jayarao, B.M., Donaldson, S.C., Straley, B.A., Sawant, A.A., Hegde, N.V., and Brown, J.L. (2006) A survey of foodborne pathogens in bulk tank milk and raw milk consumption among farm families in Pennsylvania. J Dairy Sci 89: 2451-2458.
Jolley, K.A., and Maiden, M.C.J. (2010) BIGSdb: scalable analysis of bacterial genome variation at the population level. BMC Bioinformatics 11: 595.
Keys, A.L., Dailey, R.C., Hitchins, A.D., and Smiley, R.D. (2013) Postenrichment population differentials using buffered Listeria enrichment broth: implications of the presence of Listeria innocua on Listeria monocytogenes in food test samples. J Food Prot 76: 1854-1862.
Kwong, J.C., Mercoulia, K., Tomita, T., Easton, M., Li, H.Y., Bulach, D.M., et al. (2016) Prospective whole-genome sequencing enhances national surveillance of listeria monocytogenes. J Clin Microbiol 54: 333-342.
Latorre, A.A., Pradhan, A.K., Van Kessel, J.A.S., Karns, J.S., Boor, K.J., Rice, D.H., et al. (2011) Quantitative risk assessment of listeriosis due to consumption of raw milk. J Food Prot 74: 1268-1281.
Latorre, A.A., Van Kessel, J.S., Karns, J.S., Zurakowski, M.J., Pradhan, A.K., Boor, K.J., et al. (2010) Biofilm in milking equipment on a dairy farm as a potential source of bulk tank milk contamination with Listeria monocytogenes. J Dairy Sci 93: 2792-2802.
Letunic, I., and Bork, P. (2021) Interactive tree of life (iTOL) v5: an online tool for phylogenetic tree display and annotation. Nucleic Acids Res 49: W293-W296.
Linnan, M.J., Mascola, L., Lou, X.D., Goulet, V., May, S., Salminen, C., et al. (1988) Epidemic listeriosis associated with Mexican-style cheese. N Engl J Med 319: 823-828.
MacDonald, P.D.M., Whitwam, R.E., Boggs, J.D., MacCormack, J.N., Anderson, K.L., Reardon, J.W., et al. (2005) Outbreak of listeriosis among Mexican immigrants as a result of consumption of illicitly produced Mexican-style cheese. Clin Infect Dis 40: 677-682.
Maury, M.M., Bracq-Dieye, H., Huang, L., Vales, G., Lavina, M., Thouvenot, P., et al. (2019) Hypervirulent Listeria monocytogenes clones' adaption to mammalian gut accounts for their association with dairy products. Nat Commun 10: 2488.
Maury, M.M., Tsai, Y.H., Charlier, C., Touchon, M., Chenal-Francisque, V., Leclercq, A., et al. (2016) Uncovering Listeria monocytogenes hypervirulence by harnessing its biodiversity. Nat Genet 48: 308-313.
Mohammed, H.O., Stipetic, K., McDonough, P.L., Gonzalez, R.N., Nydam, D.V., and Atwill, E.R. (2009) Identification of potential on-farm sources of Listeria monocytogenes in herds of dairy cattle. Am J Vet Res 70: 383-388.
Moura, A., Criscuolo, A., Pouseele, H., Maury, M.M., Leclercq, A., Tarr, C., et al. (2016) Whole genome-based population biology and epidemiological surveillance of Listeria monocytogenes. Nat Microbiol 2: 16185.
Moura, A., Lefrancq, N., Wirth, T., Leclercq, A., Borges, V., Gilpin, B., et al. (2021) Emergence and global spread of Listeria monocytogenes main clinical clonal complex. Sci Adv 7. https://pubmed.ncbi.nlm.nih.gov/34851675/
Moura, A., Tourdjman, M., Leclercq, A., Hamelin, E., Laurent, E., Fredriksen, N., et al. (2017) Real-time whole-genome sequencing for surveillance of Listeria monocytogenes, France. Emerg Infect Dis 23: 1462-1470.
Murinda, S.E., Nguyen, L.T., Nam, H.M., Almeida, R.A., Headrick, S.J., and Oliver, S.P. (2004) Detection of sorbitol-negative and sorbitol-positive Shiga toxin-producing Escherichia coli, Listeria monocytogenes, Campylobacter jejuni, and Salmonella spp. in dairy farm environmental samples. Foodborne Pathog Dis 1: 97-104.
Nightingale, K.K., Schukken, Y.H., Nightingale, C.R., Fortes, E.D., Ho, A.J., Her, Z., et al. (2004) Ecology and transmission of Listena monocytogenes infecting ruminants and in the farm environment. Appl Environ Microbiol 70: 4458-4467.
Oliver, S.P., Jayarao, B.M., and Almeida, R.A. (2005) Foodborne pathogens in milk and the dairy farm environment: food safety and public health implications. Foodborne Pathog Dis 2: 115-129.
Orsi, R.H., and Wiedmann, M. (2016) Characteristics and distribution of Listeria spp., including Listeria species newly described since 2009. Appl Microbiol Biotechnol 100: 5273-5287.
Painset, A., Björkman, J.T., Kiil, K., Guillier, L., Mariet, J.-F., Félix, B., et al. (2019) LiSEQ - whole-genome sequencing of a cross-sectional survey of Listeria monocytogenes in ready-to-eat foods and human clinical cases in Europe. Microb Genomics 5: e000257.
Palacios-Gorba, C., Moura, A., Leclercq, A., Gómez-Martín, Á., Gomis, J., Jiménez-Trigos, E., et al. (2021) Listeria spp. isolated from tonsils of wild deer and boars: genomic characterization. Appl Environ Microbiol 87: e02651-20.
Papić, B., Pate, M., Félix, B., and Kušar, D. (2019) Genetic diversity of Listeria monocytogenes strains in ruminant abortion and rhombencephalitis cases in comparison with the natural environment. BMC Microbiol 19: 299.
Pérez-Trallero, E., Zigorraga, C., Artieda, J., Alkorta, M., and Marimón, J.M. (2014) Two outbreaks of Listeria monocytogenes infection, northern Spain. Emerg Infect Dis 20: 2155-2157.
Prjibelski, A., Antipov, D., Meleshko, D., Lapidus, A., and Korobeynikov, A. (2020) Using SPAdes de novo assembler. Curr Protoc Bioinformatics 70: e102.
Quereda, J.J., Leclercq, A., Moura, A., Vales, G., Gómez-Martín, Á., García-Muñoz, Á., et al. (2020) Listeria valentina sp. nov., isolated from a water trough and the faeces of healthy sheep. Int J Syst Evol Microbiol 70: 5868-5879.
Ragon, M., Wirth, T., Hollandt, F., Lavenir, R., Lecuit, M., Le Monnier, A., and Brisse, S. (2008) A new perspective on Listeria monocytogenes evolution. PLoS Pathog 4: e1000146.
Ramage, C.P., Low, J.C., McLauchlin, J., and Donachie, W. (1999) Characterisation of listeria ivanovii isolates from the UKusing pulsed-field gel electrophoresis. FEMS Microbiol Lett 170: 349-353.
Reneau, J.K., Seykora, A.J., Heins, B.J., Endres, M.I., Farnsworth, R.J., and Bey, R.F. (2005) Association between hygiene scores and somatic cell scores in dairy cattle. J Am Vet Med Assoc 227: 1297-1301.
Robertson, J., and Nash, J.H.E. (2018) MOB-suite: software tools for clustering, reconstruction and typing of plasmids from draft assemblies. Microb Genomics 4: e000206.
Roche, J.R., Friggens, N.C., Kay, J.K., Fisher, M.W., Stafford, K.J., and Berry, D.P. (2009) Invited review: body condition score and its association with dairy cow productivity, health, and welfare. J Dairy Sci 92: 5769-5801.
Sanaa, M., Poutrel, B., Menard, J.L., and Serieys, F. (1993) Risk factors associated with contamination of raw milk by Listeria monocytogenes in dairy farms. J Dairy Sci 76: 2891-2898.
Sauders, B.D., Overdevest, J., Fortes, E., Windham, K., Schukken, Y., Lembo, A., and Wiedmann, M. (2012) Diversity of Listeria species in urban and natural environments. Appl Environ Microbiol 78: 4420-4433.
Schlech, W.F., Lavigne, P.M., Bortolussi, R.A., Allen, A.C., Haldane, E.V., Wort, A.J., et al. (1983) Epidemic listeriosis - evidence for transmission by food. N Engl J Med 308: 203-206.
Skovgaard, N., and Morgen, C.A. (1988) Detection of Listeria spp. in faeces from animals, in feeds, and in raw foods of animal origin. Int J Food Microbiol 6: 229-242.
Stasiewicz, M.J., Oliver, H.F., Wiedmann, M., and den Bakker, H.C. (2015) Whole-genome sequencing allows for improved identification of persistent Listeria monocytogenes in food-associated environments. Appl Environ Microbiol 81: 6024-6037.
Swaminathan, B., and Gerner-Smidt, P. (2007) The epidemiology of human listeriosis. Microbes Infect 9: 1236-1243.
Thouvenot, P., Vales, G., Bracq-Dieye, H., Tessaud-Rita, N., Maury, M.M., Moura, A., et al. (2018) MALDI-TOF mass spectrometry-based identification of Listeria species in surveillance: a prospective study. J Microbiol Methods 144: 29-32.
Van Kessel, J.A.S., Karns, J.S., Lombard, J.E., and Kopral, C.A. (2011) Prevalence of Salmonella enterica, Listeria monocytogenes, and Escherichia coli virulence factors in bulk tank milk and in-line filters from U.S. dairies. J Food Prot 74: 759-768.
Vázquez-Boland, J.A., Kuhn, M., Berche, P., Chakraborty, T., Domı́nguez-Bernal, G., Goebel, W., et al. (2001) Listeria pathogenesis and molecular virulence determinants. Clin Microbiol Rev 14: 584-640.
Vilar, M.J., Yus, E., Sanjuán, M.L., Diéguez, F.J., and Rodríguez-Otero, J.L. (2007) Prevalence of and risk factors for Listeria species on dairy farms. J Dairy Sci 90: 5083-5088.
Walland, J., Lauper, J., Frey, J., Imhof, R., Stephan, R., Seuberlich, T., and Oevermann, A. (2015) Listeria monocytogenes infection in ruminants: is there a link to the environment, food and human health? A review. Schweiz Arch Tierheilkd 157: 319-328.
Wang, H., Luo, L., Zhang, Z., Deng, J., Wang, Y., Miao, Y., et al. (2018) Prevalence and molecular characteristics of Listeria monocytogenes in cooked products and its comparison with isolates from listeriosis cases. Front Med 12: 104-112.
Wiedmann, M., Bruce, J.L., Keating, C., Johnson, A.E., McDonough, P.L., and Batt, C.A. (1997) Ribotypes and virulence gene polymorphisms suggest three distinct Listeria monocytogenes lineages with differences in pathogenic potential. Infect Immun 65: 2707-2716.
Winter, P., Schilcher, F., Bagò, Z., Schoder, D., Egerbacher, M., Baumgartner, W., and Wagner, M. (2004) Clinical and histopathological aspects of naturally occurring mastitis caused by Listeria monocytogenes in cattle and ewes. J Vet Med Ser B Infect Dis Vet Public Heal 51: 176-179.
Yusuf, S., Piedimonte, G., Auais, A., Demmler, G., Krishnan, S., Van Caeseele, P., et al. (2007) The relationship of meteorological conditions to the epidemic activity of respiratory syncytial virus. Epidemiol Infect 135: 1077-1090.
Zhang, H., Chen, W., Wang, J., Xu, B., Liu, H., Dong, Q., and Zhang, X. (2020) 10-year molecular surveillance of Listeria monocytogenes using whole-genome sequencing in Shanghai, China, 2009-2019. Front Microbiol 11: 551020.
Zhao, Q., Hu, P., Li, Q., Zhang, S., Li, H., Chang, J., et al. (2021) Prevalence and transmission characteristics of Listeria species from ruminants in farm and slaughtering environments in China. Emerg Microbes Infect 10: 356-364.
Zundel, E., and Bernard, S. (2006) Listeria monocytogenes translocates throughout the digestive tract in asymptomatic sheep. J Med Microbiol 55: 1717-1723.

Auteurs

Carla Palacios-Gorba (C)

Departamento Producción y Sanidad Animal, Salud Pública Veterinaria y Ciencia y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Cardenal Herrera-CEU, CEU Universities, Valencia, Spain.

Alexandra Moura (A)

Institut Pasteur, National Reference Centre and WHO Collaborating Centre for Listeria, Paris, France.
Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, 75015, France.

Jesús Gomis (J)

Departamento Producción y Sanidad Animal, Salud Pública Veterinaria y Ciencia y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Cardenal Herrera-CEU, CEU Universities, Valencia, Spain.

Alexandre Leclercq (A)

Institut Pasteur, National Reference Centre and WHO Collaborating Centre for Listeria, Paris, France.
Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, 75015, France.

Ángel Gómez-Martín (Á)

Departamento Producción y Sanidad Animal, Salud Pública Veterinaria y Ciencia y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Cardenal Herrera-CEU, CEU Universities, Valencia, Spain.

Hélène Bracq-Dieye (H)

Institut Pasteur, National Reference Centre and WHO Collaborating Centre for Listeria, Paris, France.
Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, 75015, France.

María L Mocé (ML)

Departamento Producción y Sanidad Animal, Salud Pública Veterinaria y Ciencia y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Cardenal Herrera-CEU, CEU Universities, Valencia, Spain.

Nathalie Tessaud-Rita (N)

Institut Pasteur, National Reference Centre and WHO Collaborating Centre for Listeria, Paris, France.
Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, 75015, France.

Estrella Jiménez-Trigos (E)

Departamento Producción y Sanidad Animal, Salud Pública Veterinaria y Ciencia y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Cardenal Herrera-CEU, CEU Universities, Valencia, Spain.

Guillaume Vales (G)

Institut Pasteur, National Reference Centre and WHO Collaborating Centre for Listeria, Paris, France.
Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, 75015, France.

Ángel García-Muñoz (Á)

Departamento Producción y Sanidad Animal, Salud Pública Veterinaria y Ciencia y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Cardenal Herrera-CEU, CEU Universities, Valencia, Spain.

Pierre Thouvenot (P)

Institut Pasteur, National Reference Centre and WHO Collaborating Centre for Listeria, Paris, France.
Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, 75015, France.

Empar García-Roselló (E)

Departamento Medicina y Cirugía Animal, Facultad de Veterinaria, Universidad Cardenal Herrera-CEU, CEU Universities, Valencia, Spain.

Marc Lecuit (M)

Institut Pasteur, National Reference Centre and WHO Collaborating Centre for Listeria, Paris, France.
Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, 75015, France.
Necker-Enfants Malades University Hospital, Division of Infectious Diseases and Tropical Medicine, Institut Imagine, APHP, Paris, France.

Juan J Quereda (JJ)

Departamento Producción y Sanidad Animal, Salud Pública Veterinaria y Ciencia y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Cardenal Herrera-CEU, CEU Universities, Valencia, Spain.

Articles similaires

Smoking Cessation and Incident Cardiovascular Disease.

Jun Hwan Cho, Seung Yong Shin, Hoseob Kim et al.
1.00
Humans Male Smoking Cessation Cardiovascular Diseases Female
Humans United States Aged Cross-Sectional Studies Medicare Part C
1.00
Humans Yoga Low Back Pain Female Male
Humans Meals Time Factors Female Adult

Classifications MeSH