| Objective:Microbial resistance is a major global public health safety concern,and antibiotic abuse is common in drug-abusing populations,and the health risks associated with it should be taken seriously.The aim of this study was to investigate the use and perception of antibiotics in the drug-using,methadone maintenance,compulsory isolation and healthy populations in Mengzi,Yunnan,and to understand the risk of antibiotic abuse in these populations.The study also examined the distribution of antibiotic resistance genes and mobile genetic elements in the intestinal tract of the drug-abusing,methadone maintenance,compulsory isolation and healthy populations in Mengzi,Yunnan,and analysed the composition and structure of the intestinal microorganisms and their association with drug resistance genes in combination with the sequencing results of intestinal microorganisms in the target population.The aim was to provide a data base for further analysis of the causes of drug resistance,to identify highly resistant flora and to guide the correct use of antibiotics in the target population.Methods:The study area was selected from Mengzi City,Honghe Prefecture,Yunnan Province,and 444 individuals were screened for drug use,methadone maintenance treatment,compulsory isolation,and healthy control groups.The genomic DNA was extracted from the faecal samples of the target population and analysed by 16S r DNA sequencing to determine the diversity and structural differences of their intestinal flora.Based on the Wafer Gen Smart Chip q PCR platform,the distribution of 35common antibiotic resistance genes and mobile genetic elements in different populations,including tetracyclines,aminoglycosides,β-lactams,mucomycins,quinolones,sulfonamides,vancomycins,MLSBs(macrolides-lincosamides-streptogrami-ns B)and other classes,were detected.The distribution characteristics of the resistance genes and their association with the intestinal flora were explored in relation to the gut microbial composition,structure and diversity of the target population.Results:(1)General demographic information showed that the four cohorts were not significantly different in terms of gender(X~2=88.797,P<0.01),age(X~2=181.676,P<0.01),education(X~2=171.010,P<0.01),marital status(X~2=115.303,P<0.01),household size(X~2=151.434,P<0.01)and annual household income(X~2=177.911,P<0.01)were all statistically significant differences.The compulsory isolation group scored significantly lower than the remaining three groups in terms of antibiotic awareness score levels(F=70.156,P<0.001).The difference in scores in the level of antibiotic use behavioural attitudes among the four groups was statistically significant(F=43.338,P<0.001).A univariate analysis of demographic characteristics of the antibiotic use behaviour scores for all groups found that differences in gender(t=-4.941,P<0.001),ethnicity(F=9.269,P<0.001),education(F=34.720,P<0.001),marital status(F=7.69,P<0.001)and household size(F=4.643,P<0.001)were all statistically significant.Among them,the level of antibiotic cognitive score and the level of antibiotic use behavioural score in the compulsory isolation group were significantly lower than those in the remaining three groups,and there is an urgent need for multi-method interventions to improve the level of cognition and safeguard health.(2)The study found differences in the distribution of gut microbial resistance genes among the drug-using,methadone maintenance treatment,compulsory isolation and healthy groups.Six drug resistance genotypes were detected in the four populations,includingβ-lactams,tetracyclines,sulfonamides,MLSBs,mucormycins and quinolones,including CMY-4、CMY-111、CTX-M-15、CTX-M-24、NDM-5、SHV-1、TEM-1、TEM-156、tet M-01、tet M-02、dfr A1、dfr A5、dfr A12、dfr A15、dfr A17、dfr A25、erm B、mcr-1and qnr B and 19 drug-resistant genotypes,of whichβ-lactams and tetracyclines were the main resistant genotypes(66.4%).Further comparative analysis revealed that the Shannon index and abundance of antibiotic resistance genes(ARGs)were significantly higher in the drug use group than in the other three groups(P<0.05),containing more types and numbers of resistance genes,and the mucormycin resistance genes were more abundant.When the four groups were compared between the two groups,the relative abundance of MLSB-like ARGs was found to be significantly higher in the MLSB group than in the remaining three groups(P<0.05),while the difference between the remaining three groups was not significant(P>0.05).The relative abundance of Colistin-like ARGs was significantly higher in the drug use group than in the other three groups(P<0.05).erm B,tet M-01,tet M-02,TEM-1 and TEM-156 were significantly enriched in all four groups.The abundance of erm B was significantly higher in the compulsory isolation group than in the other groups(P<0.05).In contrast,dfr A1 abundance was significantly higher in the drug use group than in the healthy control group(P<0.01),and mcr-1 levels were significantly more abundant than in the compulsory isolation and methadone maintenance treatment groups(P<0.01).Ten MGEs,including integrons and transposons,were detected,and the Shannon index and total abundance of MGEs in the drug-addicted group were significantly higher than those in the other three groups(P<0.05),and were significantly enriched in tnp A-01,suggesting a higher risk of drug-resistant gene transmission in the drug-addicted group.(3)The results of the gut microbial analysis showed that the diversity and structure of the gut microorganisms in the drug-using population,the methadone maintenance treatment population,the compulsory isolation population and the healthy population were different.Alpha diversity analysis of gut microorganisms revealed that the Chao1 and ACE indices were significantly higher in the methadone maintenance treatment population than in the healthy control population(P<0.05).Further PERMANOVA analysis revealed significant differences in gut microbial community structure between the four cohorts(P<0.01).Analysis of the community composition revealed that at the phylum level,the phylum Bacteroidetes(55.17%),the phylum Thick-walled(33.81%)and the phylum Aspergillus(8.97%)were the three main dominant phyla.At the genus level,Bifidobacterium,Lactobacillus,Streptococcus and Acinetobacter,Intestinibacter,Fusicatenibacter and Atopobium were significantly higher in relative abundance in the methadone maintenance group than in the remaining three groups(P<0.05).The relative abundance of Clostridium＿sensu＿stricto,Haemophilus and Roseburia was significantly higher in the drug-treatment group than in the other three groups(P<0.05).In addition,the relative abundance of Enterobacteriaceae in the healthy control group was significantly lower than that in the methadone maintenance treatment group and the compulsory isolation group(P<0.05).(4)Further correlation analysis pointed out that gut microbial diversity and composition were significantly associated with ARGs.Among them,Simpson’s index was significantly and positively correlated with dfr A12,tet M-01,tet M-02 and qnr B(r>0.3,P<0.05).At the genus level,Escherichia spp.and Shigella spp.were significantly positively correlated with TEM-1 and TEM-156(r=0.68,P<0.001),Synechococcus was significantly positively correlated with CTX-M-15(r=0.51,P<0.001),Citrobacter spp.was significantly positively correlated with dfr A12(r=0.54,P<0.001),and Stomatobaculum spp.were significantly positively correlated with SHV-1(r=0.51,P<0.001).Further potential hosts for drug-resistant genes were indicated,suggesting health risks.Conclusions:(1)The drug use and drug treatment groups were predominantly male,low-educated,unmarried and divorced and low-income.The knowledge of antibiotic use and modes of resistance transmission in this group was extremely low,making it highly susceptible to antibiotic abuse and widespread spread of antibiotic resistance genes.(2)All four groups of people were most abundant inβ-lactams,tetracyclines,mucormycins and MLSBs,and the number and types of ARGs and MGEs were most abundant in the drug use group.(3)The number of ARGs and Mobile Genetic Elements(MGEs),Shannon index and abundance were significantly higher in the drug use group than in the remaining three groups,and their scores on antibiotic awareness and behavioural attitudes towards use were also relatively lower.(4)The relative abundance of mucormycin-like resistance genes was significantly higher in the drug use group than in the remaining three groups,and the abundance of MLSB-like resistance genes was significantly higher in the compulsory isolation group than in the remaining three groups.(5)The ARG profile was significantly correlated with microbial OTUs,which may help to screen for gut microorganisms with potential risk of drug resistance gene transmission. |
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