Bacterial community diversity in Dermatophagoides farinae using high-throughput sequencing / 中国血吸虫病防治杂志

Objective To investigate the bacterial community diversity in Dermatophagoides farinae. Methods Laboratory-cultured D. farinae was collected, and the composition of microbial communities was determined by sequence analyses of the V4 region in the bacterial 16S ribosomal RNA (16S rRNA) gene on an Illumina PE250 high-throughput sequencing platform. Following quality control and filtering of the raw sequence files, valid reads were obtained and subjected to operational taxonomic units (OTU) clustering and analysis of the composition of microbial communities and alpha diversity index using the Usearch software, Silva database, and Mothur software. Results A total of 187 616 valid reads were obtained, and 469 OTUs were clustered based on a sequence similarity of more than 97%. OTU annotation showed that the bacteria in D. farinae belonged to 26 phyla, 43 classes, 100 orders, 167 families and 284 genera. The bacteria in D. farinae were mainly annotated to five phyla of Proteobacteria, Firmicutes, Bacteroidota, Actinobacteriota, and Acidobacteriota, with Proteobacteria as the dominant phylum, and mainly annotated to five dominant genera of Ralstonia, norank-f-Mitochondria, Staphylococcus and Sphingomonas, with Wolbachia identified in the non-dominant genus. Conclusions A high diversity is identified in the composition of the bacterial community in D. farinae, and there are differences in bacterial community diversity and abundance among D. farinae.

DNA barcoding of 4 species of cheyletid mites based on COI and 18S rRNA gene sequences / 中国血吸虫病防治杂志

Objective To analyze the sequences of mitochondrial cytochrome C oxidase subunit I gene (COI) and 18S ribosomal RNA gene (18S rRNA), so as to identify the feasible DNA barcodes for 4 species of cheyletid mites and improve the DNA barcoding database for cheyletid mites. Methods Cheyletid mite samples were collected from small-scale flour mills in Fuyang, Wuhu and Tongling cities of Anhui Province from May 2018 to July 2019, extracted and morphologically identified. Then, genomic DNA was extracted from a single cheyletid mite, and the COI and 18S rRNA gene sequences were obtained by PCR amplification, cloning and sequencing. The obtained sequences were aligned using the BLAST software. Multiple sequence alignment was done using the software ClustalX version 1.83 using the known gene sequences from cheyletid mites. The genetic distance was calculated using the software MEGA X, and the phylogenetic tree was created using the maximum likelihood method. Results The DNA barcoding results of Cheyletus malaccensis, C. carnifex and Cheletomorpha lepidopterorum were consistent with the morphological identification, while no sequences pertaining to Eucheyletia reticulate were retrieved in the GenBank database. The proportions of A + T were 69.6% and 55.1% in the COI and 18S rRNA sequences of 4 cheyletid mites species, respectively, and the numbers of base substitutions were 137 and 46, respectively. There were 154 to 321 and 58 to 99 inter-species variation loci in the COI and 18S rRNA gene sequences of 4 cheyletid mites species, respectively, and the intra-species genetic distance was all 0.020 or less in the COI and 18S rRNA gene sequences of 4 cheyletid mites species, with inter-species genetic distance of 0.235 to 0.583 and 0.078 to 0.114, respectively. Phylogenetic analysis based on COI and 18S rRNA genes showed that all four species of cheyletid mites were clustered into a branch with a 100% supportive rate, which was consistent with the morphological identification. Conclusion Mitochondrial COI gene is superior to 18S rRNA gene as DNA barcodes for 4 species of cheyletid mites, which is more suitable to be used to investigate the phylogenetic relationship of at genus and species levels.

[Human skin infestation caused by Haemolaelaps casalis: a case report].

Our study reports a case of acarodermatitis caused by Haemolaelaps casalis. By morphological observations, the mites seized were identified as Haemolaelaps casalis (deutonymph) which could attack humans resulting in acarodermatitis characterized with the symptoms of papules and blisters in different degrees. The patient was treated with 15% calamine lotion and anti-inflammatory and antipruritic drugs. Meanwhile, the mites were eliminated in the bedroom. After the treatment for one week, the patient was cured. Haemolaelaps casalis, which had been found in the indoor mattress, could attack humans and cause acarodermatitis. We should strengthen the work of anti-mite in domestic environment.

Human skin infestation caused by Haemolaelaps casalis: a case report / 中国血吸虫病防治杂志

Our study reports a case of acarodermatitis caused by Haemolaelaps casalis. By morphological observations, the mites seized were identified as Haemolaelaps casalis (deutonymph) which could attack humans resulting in acarodermatitis characterized with the symptoms of papules and blisters in different degrees. The patient was treated with 15% calamine lotion and anti-inflammatory and antipruritic drugs. Meanwhile, the mites were eliminated in the bedroom. After the treatment for one week, the patient was cured. Haemolaelaps casalis, which had been found in the indoor mattress, could attack humans and cause acarodermatitis. We should strengthen the work of anti-mite in domestic environment.

[Optimization and comparison of extraction methods of mitochondrial DNA of Oncomelania hupensis].

OBJECTIVE: To optimize the extraction methods of mitochondrial genome DNA (mtDNA) of Oncomelania hupen- sis. METHODS: The pyrolysis, protein K variable-temperature digestion and high-concentration potassium acetate purification were applied to optimize the high-concentration-salt precipitation method, and then the optimized method was compared with two common extraction methods, the sucrose density gradient centrifugation method and traditional high-concentration-salt precipitation method. The mtDNA samples were identified by using spectrophotometry, agarose gel electrophoresis and the amplification products of COX1. The nuclear DNA contamination was tested by the amplification products of ITS. RESULTS: The concentration and yield of the improved method was significantly higher than those of the traditional method (F = 3 032.65, 10 185.00, both P < 0.01). The mtDNA samples extracted were essentially free of nuclear DNA and protein, meeting PCR, sequence analysis and other molecular biology research requirements. CONCLUSIONS: The improved high-concentration-salt precipitation method for isolating mtDNA is simple, and it has high yield and low cost. The extracted mtDNA can meet relevant analysis requirements.

[Study on natural population dynamics and spatial distribution pattern of Tyrophagus putrescentiae in stored flour].

OBJECTIVE: To understand the natural population dynamics and spatial distribution of Tyrophagus putrescentiae in storage flour, so as to provide an evidence for its prevention and control. METHODS: The samples from five sampling points in Wuhu City were collected monthly from January to December, 2013, and examined and counted for T. putrescentiae. The dispersion pattern target, Iwao's m*- x̄; regression analysis and Taylor's lgS2-lg x̄; regression analysis were used for analyzing the spatial distribution pattern of T. putrescentiae in the storage flour. RESULTS: The peaks of population dynamics of T. putrescentiae were discovered in July and September. The indexes of dispersion were as follows：I > 0, CA > 0, m*/ x̄; > 1; and the linear regression equation of Iwao：m* = 3.740 3 + 1.017 5 x̄; (r = 0.995 8) and Taylor：lgS2 = 0.500 4 + 1.134 9 lg x̄; (r =0.832 8) showed that the spatial distribution pattern of T. putrescentiae in the storage flour was assembled. CONCLUSIONS: The peak of population dynamics of T. putrescentiae in the storage flour in Wuhu City is a double peak type, and the spatial distribution pattern of T. putrescentiae is assembled.

[Comparison of three different methods for isolating RNA from Oncomelania hupensis].

OBJECTIVE: To compare the effects of three kinds of Oncomelania hupensis RNA extraction methods, namely a modified SDS method, TRIzol reagent method, and CTAB method, so as to obtain an economical and efficient method for RNA extraction from O. hupensis. METHODS: The modified SDS method, TRIzol reagent method and CTAB method were applied to extract the RNA from O. hupensis. A nucleic acid protein analyzer was used to measure the concentration and purity of RNA. The yields were calculated by the concentration of the products. The purity was indicated by A260/A280 and A260/A230. The quality of RNA was inspected by 1% agarose gel electrophoresis. The ß-acting gene was selected as the target gene for RT-PCR analysis. RESULTS: The RNA yields obtained by using the three kinds of extraction methods were significantly different (F = 16 895.85, P < 0.01) according to the analysis of variance. The LSD test showed that the yields obtained by using the modified SDS method were the highest, and those obtained by the CTAB method were the lowest. The purity of RNA extracted by the CTAB method was superior to that by the other two methods, and the A260/A280 and A260/A230 ratios of the CTAB method were in the range from 1.8-2.0 and 2.0-2.2. The A260/A230 ratios of the other two methods were both lower than 2.0. The RNA extracted by the modified SDS method had the better integrity. The electrophoresis results showed that the 28S rRNA band, 18S rRNA band and 5S rRNA band were clear, and there was no obvious smear between each band. The RNA obtained by the TRIzol reagent method had no 28S rRNA band, and that obtained by the CTAB method had no 28S rRNA and 5S rRNA bands. The ß-acting gene of the RNA extracted by all the three methods could be amplified by RT-PCR. The costs and time-consuming of the modified SDS method were less than those of the other two methods. CONCLUSIONS: The modified SDS method is an economic and efficient method, and it is suitable for extracting the RNA of O. hupensis, especially for large sample preparation.

[Investigation on mites and insects breeding status of bed mats in college dormitories in Wuhu City].

OBJECTIVE: To understand the breeding situations of mites and insects from bed mats in dormitories in a college in Wuhu City, so as to provide evidences for improving the prevention and control of mites. METHODS: From March to May, 2015, the dust samples from bed mats in student dormitories were collected and detected for mites and insects by microscopy. In addition, the intervention measures including soaking the mats in the warm water, exposing the mats to the sunlight for a long time and cleaning up the indoor environment were carried out, and the breeding situations of mites and insects before and after the intervention were compared. RESULTS: A total of 428 dust samples from bed mats were collected, and the total infestation rate of mites and insects was 71.03%. There were 11 species of mites and insects identified, among which, the infestation rate of Dermatophagoides farinae (60.05%) was the highest, following by that of Liposcelis bostrychophilus (40.89%) . The infestation rates of Cheyletus malaccensis, Blattisocius sp. and Pyemotes sp. were 9.81%, 3.74% and 1.64%, respectively, and the above mites might cause dermatits. The infestation rates of mites and insects in dust samples from bed mats stored in indoor corner, wardrobe, bed bottom, and the balcony corner were 74.75%, 71.26%, 61.17%, and 77.78%, respectively, and the differences among them were not statistically significant (χ2 = 7.030, P > 0.05) . The detectable rates of mites and insects in dust samples from bed mats with no cover, wrapped with cloth bags, and wrapped with plastic bags were 85.58%, 78.13%, and 14.29%, respectively, and the differences among them were statistically significant (χ2 = 164.303, P < 0.05) . After the intervention, both the infestation rates of mites and insects as well as the average density of mites were decreased, and the differences were statistically significant (χ2 = 45.615, t = 3.203, both P < 0.05) . CONCLUSIONS: The infestation rates of mites and insects in bed mats of the dormitories in the college of Wuhu City are high, and among all the species of mites infested, D. farina are preponderant. The intervention measures, such as prepacking bed mats by adequate sealing, soaking bed mats in the warm water, exposing the bed mats to the sunlight and cleaning indoor environment, have an inhibiting effect to the infestation of mites and insects.

[Tyrophagus palmarum and its hypopus found in feed of Tenebrio molitor].

OBJECTIVE: To observe the Tyrophagus palmarum and its hypopus in feed of Tenebrio molitor. METHODS: Feed samples were collected from T. molitor farms, and T. palmarum and its hypopus were separated from the samples by the vibration sieve method and floating method. The glass specimens were prepared and T. palmarum and its hypopus were observed under an optical microscope. RESULTS: Under the light microscope, the skin of T. palmarum was smooth and bright, the color of legs and chelicerae was relatively dark, the length of dorsal seta d2 was 3-4 times longer than d1 and la; The ω1 in tarsus of leg Ⅰ, Ⅱ was short rod, and its top was not tapered; the sucker plate in tarsus of leg Ⅳ located in the middle of the section. The hypopus was oblate, and there were two pairs of legs in front which extended to body. The rear end of hypopus became pointless and round, wrapping around a transparent and ossification skin. CONCLUSIONS: Light microscopy shows the morphological characteristics of T. palmarum and its dormant body, providing the basis for identifying the hypopus.

[Population dynamics and spatial distribution pattern of Aleuroglyphus ovatus].

OBJECTIVE: To characterize the population dynamics and spatial distribution of Aleuroglyphus ovatus in the flour warehouse, so as to provide the basic evidence for improving the sampling guidelines that are essential for effective pest monitoring and management. METHODS: The samples from flour warehouses of four localities were collected, examined and counted for A. ovatus in every month in Wuhu City. The dispersion pattern target, Iwao m*/x̅ regression analysis and Taylor power method were used for analyzing the spatial distribution pattern of A. ovatus in the flour warehouses. RESULTS: The peaks of population dynamics of A. ovatus were discovered in July and September, respectively. The indexes of dispersion were as follows：I > 0, CA > 0, m*/x̅ > 1. At the same time, the parameters in the equation of Iwao：m*=5.471+1.022 x̅ (r = 0.999) and Taylor： lgS2 = 0.697+1.111 lg x̅ (r = 0.987) showed that the spatial distribution pattern of A. ovatus was assembled. CONCLUSIONS: The peaks of population dynamics of A. ovatus in the flour warehouse are bimodal pattern, and the spatial distribution pattern of A. ovatus is assembled.