The influence of primer choice on archaeal phylogenetic analyses based on 16S rRNA gene PCR / A influência da escolha de iniciadores na filogenia de Archaea em ensaios de PCR para o gene rRNA 16S

Abstract Polymerase chain reaction (PCR) assays targeting 16S rRNA genes followed by DNA sequencing are still important tools to characterize microbial communities present in environmental samples. However, despite the crescent number of deposited archaeal DNA sequences in databases, until now we do not have a clear picture of the effectiveness and specificity of the universal primers widely used to describe archaeal communities from different natural habitats. Therefore, in this study, we compared the phylogenetic profile obtained when Cerrado lake sediment DNA samples were submitted to 16S rDNA PCR employing three Archaea-specific primer sets commonly used. Our findings reveal that specificity of primers differed depending on the source of the analyzed DNA. Furthermore, archaeal communities revealed by each primer pair varied greatly, indicating that 16S rRNA gene primer choice affects the community profile obtained, with differences in both taxon detection and operational taxonomic unit (OTU) estimates.

Resumo A amplificação de genes que codificam o rRNA 16S por reação em cadeia da polimerase (PCR) e o seu subsequente sequenciamento consistem em uma ferramenta importante na caracterização de comunidades microbianas presentes em amostras ambientais. No entanto, apesar do crescente número de sequências de DNA de Archaea depositadas em bancos de dados, a especificidade e efetividade dos iniciadores de PCR descritos como universais e amplamente utilizados na descrição desse grupo ainda não está clara. Neste estudo foram comparados os perfis filogenéticos de comunidades de arqueias obtidos a partir amostras de DNA de sedimentos lacustres do Cerrado submetidas a ensaios de PCR empregando três pares de iniciadores específicos para Archaea, comumente utilizados neste tipo de estudo. Nossos resultados indicam que as comunidades de arqueias detectadas com cada par de iniciadores apresentaram grande variação filogenética, sugerindo que a escolha de iniciadores dirigidos ao gene de rRNA 16S tem efeito significativo no perfil da comunidade descrita, com diferenças tanto em relação aos táxons detectados, como nas estimativas de unidades taxonômicas operacionais (OTU).

Bacterial community associated with Pyrodinium bahamense var. compressum during late exponential growth phase based on 16S rRNA gene sequence analysis

Aims@#Pyrodinium bahamense var. compressum is one of the principal causal agents of harmful algal blooms (HABs) in the coastal waters of Sabah, Malaysia. Seafood and aquaculture products tainted with lethal concentrations of the principal neurotoxin, saxitoxin, have been implicated in mortality and morbidity. The bacteria-algae association may play a key role in paralytic shellfish toxin (PST) production during a toxic bloom event. The production of PST during a harmful bloom is unclear and research on the bacterial diversity associated with Sabah P. bahamense is scarce. The present study examined the cultivable bacteria diversity associated with P. bahamense through 16S ribosomal RNA (rRNA) gene sequence analysis.@*Methodology and results@#The V3-V4 region of the 16S rRNA gene sequence was amplified and used to identify bacterial populations associated with P. bahamense var. compressum. A total of 62 isolates were successfully isolated, belonging to three different phyla, which were Proteobacteria; 55 (89%), Bacteroidetes; 6 (10%) and Actinobacteria; 1 (1%). Out of 55 Proteobacteria, 27 isolates were gamma-Proteobacteria (Marinobacter salsuginis) and 28 of the isolates were alpha-Proteobacteria; Mameliella atlantica (13), Roseibium denhamense (10) and Roseibium hamelinense (5). The remaining bacteria isolates from the phyla Bacteroidetes and Actinobacteria were identified as Muricauda lutimaris (6) and Micrococcus luteus (1), respectively.@*Conclusion, significance and impact of study@#The analysis of the bacterial 16S rRNA gene revealed multiple bacterial taxa associated with the toxic P. bahamense var. compressum bloom. The findings of the present work will pave the way for further studies aimed at isolating and characterizing genes involved in the saxitoxin biosynthesis in the associated bacteria.

Determining the protective effects of Ma-Mu-Ran Antidiarrheal Capsules against acute DSS-induced enteritis using 16S rRNA gene sequencing and fecal metabolomics / 中国天然药物

Ma-Mu-Ran Antidiarrheal Capsules (MMRAC) is traditional Chinese medicine that has been used to treat diarrhea caused by acute enteritis (AE) and bacillary dysentery in Xinjiang (China) for many years. However, the potential therapeutic mechanism of MMRAC for AE and its regulatory mechanism on host metabolism is unclear. This study used fecal metabolomics profiling with GC/MS and 16S rRNA gene sequencing analysis to explore the potential regulatory mechanisms of MMRAC on a dextran sulfate sodium salt (DSS)-induced mouse model of AE. Fecal metabolomics-based analyses were performed to detect the differentially expressed metabolites and metabolic pathways. The 16S rRNA gene sequencing analysis was used to assess the altered gut microbes at the genus level and for functional prediction. Moreover, Pearson correlation analysis was used to integrate differentially expressed metabolites and altered bacterial genera. The results revealed that six intestinal bacteria and seven metabolites mediated metabolic disorders (i.e., metabolism of amino acid, carbohydrate, cofactors and vitamins, and lipid) in AE mice. Besides, ten altered microbes mediated the differential expression of eight metabolites and regulated these metabolisms after MMRAC administration. Overall, these findings demonstrate that AE is associated with metabolic disorders and microbial dysbiosis. Further, we present that MMRAC exerts protective effects against AE by improving host metabolism through the intestinal flora.

Evaluation of Genetic Relationship of some Squirrelfishes through DNA Barcode

Abstract Species of the subfamily Holocentrinae, family Holocentidae, commonly called, squirrelfishes, are widely distributed from tropical to warm temperate waters. In Egypt, no data are available on genetic and evolutionary relationships of the family Holocentridae. Therefore, the study of the genetic relationship among Holocentrids species is crucial for proper management and convenient strategies. The purpose of this study was to evaluate the genetic relationship among eight species belonging to the family Holocentridae from the Mediterranean Sea and the Red Sea in Egypt using DNA barcoding. Based on this molecular marker, a phylogenetic tree was constructed for the studied Holocentrids species. 12S rRNA sequences discovered that Sargocentron caudimaculatum was clustered as closest taxa to Sargocentron spiniferum, being a sister group to each other. Also, Sargocentron punctatissimum and Sargocentron macrosquamis were more related to each other and formed a sister group. Moreover, this study discusses the building of genetic relationship among Sargocentron spinosissimum and Sargocentron macrosquamis for the first time to the other studied Sargocentrons. DNA barcoding using 12S rRNA gene provided efficient DNA barcodes for all of the studied species. The constructed phylogenetic tree based on the employed molecular marker provided the update for the barcoded Holocentridae species evolution.

Mutaciones del gen ARN ribosómico 23S de Helicobacter pylori asociadas con resistencia a claritromicina en pacientes atendidos en una unidad de endoscopia de Medellín, Colombia / Mutations in the Helicobacter pylori 23S rRNA gene associated with clarithromycin resistance in patients at an endoscopy unit in Medellín, Colombia

Resumen Introducción. La claritromicina es el antibiótico de primera línea para el tratamiento de la infección por Helicobacter pylori. La resistencia bacteriana se produce principalmente por mutaciones puntuales del gen ARN ribosómico 23S (ARNr 23S). Objetivo. Determinar la frecuencia de las mutaciones puntuales A2143G y A2142G del gen ARNr 23S asociadas con la resistencia de H. pylori a la claritromicina en muestras de pacientes con manifestaciones dispépticas en Medellín, región noroccidental de Colombia. Materiales y métodos. Se extrajo ADN a partir de muestras de biopsia gástrica obtenidas de pacientes con manifestaciones dispépticas atendidos en una unidad de endoscopia entre el 2016 y el 2017. Mediante reacción en cadena de la polimerasa (PCR), se amplificaron las regiones s y m del gen vacA y una región del gen ARNr 23S bacteriano. La presencia de las mutaciones A2142G y A2143G se determinó por la técnica de polimorfismos de longitud de fragmentos de restricción (RFLP) con las enzimas BbsI y BsaI, respectivamente. Resultados. Se encontró una prevalencia de infección de 44,2 % (175/396), según el informe de histopatología. En 143 de estas 175 muestras positivas se amplificaron las tres regiones del genoma bacteriano. Se identificaron las mutaciones A2143G y A2142G en 27 muestras (18,8 %; 27/143), la mutación más frecuente fue la A2143G (81,5 %; 22/27). Conclusiones. Hubo una gran prevalencia de mutaciones asociadas con la resistencia de H. pylori a la claritromicina en la población de estudio. Se requieren estudios adicionales para establecer la resistencia bacteriana en la población colombiana y, así, determinar los tratamientos de primera línea y de rescate.

Abstract Introduction: Clarithromycin is the first-line antibiotic for the treatment of Helicobacter pylori infection. Bacterial resistance is mainly due to the presence of specific mutations in the 23S ribosomal RNA (rRNA) gene. Objective: To determine the frequency of A2143G and A2142G specific mutations in the 23S rRNA gene associated with clarithromycin resistance of H. pylori in samples from patients with dyspeptic manifestations in Medellín, northwestern Colombia. Materials and methods: DNA was extracted from gastric biopsy samples of patients with dyspeptic manifestations seen at an endoscopy unit in Medellín between 2016 and 2017. PCR was performed to amplify the bacterial s and m vacA regions, and a region in the 23S rRNA gene. The presence of the A2142G and A2143G mutations was determined using the restriction fragment length polymorphism (RFLP) technique with the BbsI and BsaI enzymes, respectively. Results: The prevalence of infection was 44.2% (175/396), according to the histopathology report. The positive samples were analyzed and the three regions of the bacterial genome were amplified in 143 of the 175 samples. The A2143G and A2142G mutations were identified in 27 samples (18.8%, 27/143). The most frequent mutation was A2143G (81.5%, 22/27). Conclusions: We found a high prevalence of H. pylori mutations associated with clarithromycin resistance in the study population. Further studies are required to determine the bacterial resistance in the Colombian population in order to define first line and rescue treatments.

Analysis of GJB2, SLC26A4, GJB3 and 12S rRNA gene mutations among patients with nonsyndromic hearing loss from eastern Shandong / 中华医学遗传学杂志

OBJECTIVE@#To explore the characteristics of mutations of four common pathogenic genes (GJB2, SLC26A4, GJB3 and 12S rRNA) among patients with nonsyndromic hearing loss (NSHL) from eastern Shandong.@*METHODS@#Peripheral blood samples of 420 NSHL patients were collected, and a hereditary-deafness-gene microarray was used to detect GJB2 c.235delC, c.299-300delAT, c.35delG and c.176del16 mutations, GJB3 c.538C>T mutation, SLC26A4 c.2168A>G and c.IVS7-2A>G mutations, and 12S rRNA c.1555A>C and c.1494C>T mutations. For patients carrying single heterozygous mutations, the coding regions of the above genes were analyzed with Sanger sequencing.@*RESULTS@#The results of the microarray assay and Sanger sequencing showed that 84 patients (20.00%) carried GJB2 mutations, with c.235delC (16.43%) and c.299-300delAT (7.86%) being most common. Seventy-five patients (17.86%) carried SLC26A4 mutations, for which c.IVS7-2A>G accounted for 15.71%. In addition, 5.95% of patients carried 12S rRNA mutations. Only one patient was found to carried GJB3 mutation (c.538C>T).@*CONCLUSION@#Common pathogenic mutations for NSHL in eastern Shandong included GJB2 c.235delC and SLC26A4 c.IVS7-2A>G. Of note, 5.95% of patients were due to 12S rRNA m.1555A>G mutation, which gave a frequency greater than other regions of China.

Co-Infection of Scrub Typhus and Human Granulocytic Anaplasmosis in Korea, 2006

BACKGROUND: Scrub typhus, severe fever with thrombocytopenia syndrome (SFTS) and human granulocytic anaplasmosis (HGA) are important arthropod-borne infectious diseases in Korea and share a common point that they are transmitted by arthropod bites mostly during outdoor activities and there are considerable overlaps of epidemiologic and clinical features at presentation. We investigated the co-infection of these infections. METHODS: The study subjects were patients with laboratory-confirmed scrub typhus who were enrolled retrospectively in 2006. SFTS virus (SFTSV) infection was confirmed by a reverse transcriptase polymerase chain reaction (PCR) to amplify partial L segment of SFTSV for molecular diagnosis. HGA was confirmed by a nested PCR to amplify 16S rRNA gene of Anaplasma phagocytophilum. Direct sequencing of the positive PCR products was performed. Clinical features of co-infected subjects were described. RESULTS: One-hundred sixty-seven patients with scrub typhus were included in the analysis. Co-infection of A. phagocytophilum was identified in 4.2% of scrub typhus patients (7/167). The route of co-infection was uncertain. The co-infected patients had not different clinical manifestations compared to the patients with scrub typhus only. All the study subjects were negative for SFTSV. CONCLUSION: We found retrospective molecular evidence of the co-infection of scrub typhus and HGA in Korea. HGA may be more prevalent than expected and need to be considered as an important differential diagnosis of febrile patients in Korea.

Facklamia hominis Isolated from a Wound: A Case Report and Review of the Literature / 대한임상미생물학회지

Facklamia hominis is a facultative anaerobic Gram-positive coccus generally displaying weak alpha-hemolysis and negativity for catalase and oxidase. Facklamia species are part of the normal flora of the female genitourinary tract and have been reported in invasive diseases such as meningitis and infective endocarditis, albeit rarely. A 67 year-old-man presented to hospital with a tender, erythematous epidermal cyst on the right side of his upper back. Simple excision of the cyst was performed and the pus was taken with a sterile swab for culture, yielding no growth. One week later, discharge was observed in the patient's wound site and a sterile swab for culture was taken. The colonies grown were identified as F. hominis by the Vitek 2 system (bioMérieux, France), and the result was then reported to clinicians, and later confirmed by 16S rRNA gene sequencing and matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry. To the best of our knowledge, this is the first reported case of F. hominis isolation from a clinical specimen in Korea.

Application of the Whole Genome-Based Bacterial Identification System, TrueBac ID, Using Clinical Isolates That Were Not Identified With Three Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) Systems

BACKGROUND: Next-generation sequencing is increasingly used for taxonomic identification of pathogenic bacterial isolates. We evaluated the performance of a newly introduced whole genome-based bacterial identification system, TrueBac ID (ChunLab Inc., Seoul, Korea), using clinical isolates that were not identified by three matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) systems and 16S rRNA gene sequencing. METHODS: Thirty-six bacterial isolates were selected from a university-affiliated hospital and a commercial clinical laboratory. Species was identified by three MALDI-TOF MS systems: Bruker Biotyper MS (Bruker Daltonics, Billerica, MA, USA), VITEK MS (bioMérieux, Marcy l'Étoile, France), and ASTA MicroIDSys (ASTA Inc., Suwon, Korea). Whole genome sequencing was conducted using the Illumina MiSeq system (Illumina, San Diego, CA, USA), and genome-based identification was performed using the TrueBac ID cloud system (www.truebacid.com). RESULTS: TrueBac ID assigned 94% (34/36) of the isolates to known (N=25) or novel (N=4) species, genomospecies (N=3), or species group (N=2). The remaining two were identified at the genus level. CONCLUSIONS: TrueBac ID successfully identified the majority of isolates that MALDI-TOF MS failed to identify. Genome-based identification can be a useful tool in clinical laboratories, with its superior accuracy and database-driven operations.

First comprehensive report of bacteria spp. associated with cloaca of Laudakia nupta (Sauria: Agamidae) in Iran using molecular studies / 한국실험동물학회지

Iran bears a remarkable variety of reptiles. One of the lizard families occurring in Iran is the Family Agamidae which is widely are distributed throughout the old world. The large-scaled rock agamid, Laudakia nupta, is one of the well-known agamid. There are few reports of cloacal microbial on reptiles hence their function in cloacae remains unknown. Laudakia nupta usually live in rural and urban areas and close vicinity to man, they are likely to play an important role in the spread of disease that may be caused by these microorganisms and their transmission to man. Therefore, the aim of this study was to identify the bacterial flora colonizing the cloacal region of Laudakia nupta using molecular studies. The cloacal fluids were directly placed on nutrient agar (NA) plates and incubated at 25 ± 2 ℃ for 48 h. The resulting bacterial colonies were transferred to fresh nutrient agar (NA) plates for molecular studies. Twelve isolates were obtained from 17 specimens of Laudakia nupta. All bacteria isolates were identified as Bacillus subtillis (5), Bacillus cereus (4), Bacillus sp. (1), Pseudomonas putida (1), and Pseudomonas sp. (1) based on partial sequences of the 16 s rRNA gene. This is the first comprehensive report of bacteria spp. associated with cloaca of Laudakia nupta using molecular studies. In this research, we found that Laudakia nupta can be a carrier of bacteria which can transfer microorganisms to hosts.

A Case of Catabacter hongkongensis and Alistipes indistinctus Isolated from Blood Cultures of a Patient with Acute Appendicitis

Catabacter hongkongensis is an anaerobic gram-positive coccobacillus that was first isolated in Hong Kong. It is infectious and causes high mortality in patients with rare but underlying diseases. Alistipes indistinctus is an anaerobic gram-negative coccobacillus. This bacterium is a common member of the human intestinal microbiota. We report a case of C. hongkongensis and A. indistinctus isolated from blood cultures of a patient with acute appendicitis. A 35-year-old female patient with no specific medical history was admitted to the hospital due to abdominal pain, vomiting, nausea, and diarrhea experienced on the day before admission. On admission, laboratory tests revealed leukocytosis, neutropenia, and elevated C–reactive protein and procalcitonin levels. Following an abdominal computed tomography showing acute appendicitis with suspected perforation, emergency surgery was performed. Growth was observed in two anaerobic blood culture bottles after four days. After further culturing of the bacteria on Brucella Blood Agar, two types of bacteria were obtained. The two bacterial isolates, one gram-positive and one gram-negative, were unable to be identified using matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Thus, 16S rRNA gene sequence analysis was performed, resulting in identification of the bacteria as C. hongkongensis and A. indistinctus. The patient was administered antibiotics and discharged two days after surgery. Although MALDI-TOF MS enables fast and accurate identification of bacteria, C. hongkongensis and A. indistinctus were not listed in the spectral library, and 16S rRNA gene sequence analysis was useful for identifying the two bacteria.

A Case of Chryseobacterium hominis Isolated from Human Blood Drawn Through Peripherally Inserted Central Catheter

Chryseobacterium hominis is non-fermenting Gram-negative rod that was first identified as a novel species in 2007. Here, we report the first clinical case of C. hominis bacteremia, which was confirmed by MALDI-TOF MS and 16S rRNA gene sequencing. A 16-year-old boy diagnosed with acute lymphoblastic leukemia was hospitalized for three months. Two sets of blood culture test through a peripherally inserted central catheter (PICC), which was inserted a month ago, was performed when his white blood cell count declined and he had a high fever. Colonies of medium sizes that looked round, mucoid, sticky, and grayish on blood and chocolate agar plates were observed. Identification of bacteria using the VITEK MALDI-TOF MS system (BioMérieux, France) was not successful and the VITEK 2 system (BioMérieux, USA) indicated Sphingomonas paucimobilis, with a questionable level of confidence (92%). However, Microflex LT Biotyper (Bruker Daltonics, Germany) showed C. homins (log score: 1.81) and sequence of 16S rRNA showed a 100% identity with C. hominis. Piperacillin-tazobactam was administered since the isolate was susceptible to piperacillin-tazobactam but C. hominis showed growth in the next four follow-up culture of blood drawn through PICC. The fever subsided only after PICC was changed. The clinical prognosis and antimicrobial susceptibility test of C. hominis should be further studied.

Molecular Detection of Coxiella burnetii in Cattle on Ulleung Island, Korea: A Population-based Study with Four Years of Follow Up

In a population-based study with 4 years of follow up, we evaluated the prevalence of Coxiella burnetii in cattle on Ulleung Island, Korea. In this study, the rates of C. burnetii infection in cattle on Ulleung Island were determined by PCR and were found to be 0.3–1.0% in the period 2011–2014. All 17 C. burnetii partial 16S rRNA gene sequences from PCR-positive cattle were identical and 2 geographic representatives were included in our analysis. The nucleotide sequences of the 2 samples showed high (98.4–100%) identity with C. burnetii sequences obtained from the GenBank. In this long-term tracking study, the number of cattle positive for C. burnetii on Ulleung Island was low. To prevent the transmission of C. burnetii on Ulleung Island, control strategy should include biosecurity improvement in surveillance, livestock management, administering suitable tests before purchasing animals to detect C. burnetii shedders, and restricting movements between herds.

Molecular Prevalence of Cryptosporidium spp. in Breeding Kennel Dogs

Cryptosporidium is a common intestinal protozoan that can lead to diarrhea in humans and dogs. The predominant species of infection are C. hominis and C. parvum in humans, and C. canis in dogs. However, C. canis can infect immunocompromised humans. Considering the close contact with humans, dogs have the potential to be reservoirs for human cryptosporidiosis. Breeding kennels are the major supply source of puppies for pet shops. The present study is to determine the molecular prevalence and characteristics of Cryptosporidium spp. found in breeding kennel dogs. A total of 314 fecal samples were collected from young and adult dogs kept in 5 breeding kennels. A polymerase chain reaction targeting the small subunit rRNA gene was employed for the detection of Cryptosporidium spp. To determine the species, the DNA sequences were compared to GenBank data. Overall, 21.0% of the fecal samples were positive for Cryptosporidium spp. infection. Cryptosporidium spp. was detected in all 5 facilities. A sequencing analysis demonstrated that all isolates shared 99–100% similarity with C. canis. The results suggest that Cryptosporidium spp. infection is present at a high-level in breeding kennel dogs. However, because dominant species in this survey was C. canis, the importance of breeding kennel dogs as reservoirs for Cryptosporidium spp. transmission to humans is likely to be low in Japan.

Identification of Candida Species Using 26S Ribosomal RNA Gene Sequencing in Patients with Periodontitis

The infection with Candida spp. for oral cavity is being increasingly reported. However, its variations have not yet been specifically described in periodontitis. The present study was conducted to use an uniplex 26S rRNA-based amplicons to detect and discriminate Candida using only one pair of ribosomal primers. A total of 50 patients with chronic periodontitis was involved in the study. Pure Candida colonies were isolated from 23 patients and genomic DNA was extracted, and PCR was conducted. Direct DNA sequencing followed by comprehensive phylogenetic analyses were performed to confirm the identity of Candida colonies. Results indicated that the ration of Candida-infected patients was 46%, with a high prevalence of C. albicans, followed by remarkably lower ratios of C. parapsilosis, C. glabrata, C. kefyr, and C. dubliniensis respectively. Phylogenetic analyses indicated obvious discrimination amongst the analyzed Candida species as each observed species occupied a distinctive phylogenetic position. The current results reported a simple, efficient, and low-cost detection of five species of Candida without the need for other costly techniques of molecular screening. The current findings may help dentists to easily take a snapshot of the patterns of Candida infection in periodontitis cases to assess the nature and grade of infection.

Campylobacter fetus Peritonitis in a Patient with Continuous Ambulatory Peritoneal Dialysis: A First Case Report in Korea / 대한임상미생물학회지

Campylobacter fetus may cause infections such as septicemia, peritonitis, meningitis, endocarditis, septic arthritis, and cellulitis, increasing the risk of spontaneous abortion but decreasing the likelihood of gastroenteritis. We identified C. fetus from continuous ambulatory peritoneal dialysis (CAPD) fluid using 16S rRNA gene sequencing. It is significant that this is the first case report in Korea of CAPD peritonitis caused by C. fetus, which is known to be rare.

Molecular identification of coagulase-negative staphylococci by rpoB sequence typing / 대한수의학회지

Bovine mastitis (BM) has resulted in enormous economic loss in the dairy industry and coagulase-negative staphylococci (CNS) have caused subclinical BM. Although VITEK 2 GP ID card (VITEK 2) has been used for CNS identification, the probability of identification varies. The rpoB sequence typing (RSTing) method has been used for molecular diagnosis and epidemiology of bacterial infections. In this study, we undertook RSTing of CNS and compared the results with those of VITEK2 and 16S rRNA gene sequencing. As compared VITEK2, the molecular-based methods were more reliable for species identification; moreover, RSTing provided more molecular epidemiological information than that from 16S rRNA gene sequencing.

Benefits of procyanidins on gut microbiota in Bama minipigs and implications in replacing antibiotics

Several studies have reported the effect of absorption of procyanidins and their contribution to the small intestine. However, differences between dietary interventions of procyanidins and interventions via antibiotic feeding in pigs are rarely reported. Following 16S rRNA gene Illumina MiSeq sequencing, we observed that both procyanidin administration for 2 months (procyanidin-1 group) and continuous antibiotic feeding for 1 month followed by procyanidin for 1 month (procyanidin-2 group) increased the number of operational taxonomic units, as well as the Chao 1 and ACE indices, compared to those in pigs undergoing antibiotic administration for 2 months (antibiotic group). The genera Fibrobacter and Spirochaete were more abundant in the antibiotic group than in the procyanidin-1 and procyanidin-2 groups. Principal component analysis revealed clear separations among the three groups. Additionally, using the online Molecular Ecological Network Analyses pipeline, three co-occurrence networks were constructed; Lactobacillus was in a co-occurrence relationship with Trichococcus and Desulfovibrio and a co-exclusion relationship with Bacillus and Spharerochaeta. Furthermore, metabolic function analysis by phylogenetic investigation of communities by reconstruction of unobserved states demonstrated modulation of pathways involved in the metabolism of carbohydrates, amino acids, energy, and nucleotides. These data suggest that procyanidin influences the gut microbiota and the intestinal metabolic function to produce beneficial effects on metabolic homeostasis.

A Case of Biliary Sepsis by Dysgonomonas capnocytophagoides

Dysgonomonas capnocytophagoides is a gram-negative, facultatively anaerobic coccobacillus that was formerly designated CDC group dysgonic fermenter (DF)-3, occurring as a normal flora in human gut and rarely causing human infections such as bacteremia, abscess, diarrhea, and cholecystitis. In this study, we report a case of biliary sepsis caused by D. capnocytophagoides in a patient with biliary obstruction. A seventy four-year-old man, admitted to the hospital due to common bile-duct stone, also had cholangitis caused by D. capnocytophagoides and Enterococcus avium, which were isolated from his blood cultures. D. capnocytophagoides was initially identified as D. gadei by MALDI-TOF mass spectrometry, but later confirmed as D. capnocytophagoides by 16S rRNA gene sequencing. To the best of our knowledge, this is the first report of human infection by D. capnocytophagoides in Korea.

Hypolobocera guayaquilensis (Decapoda: Pseudothelphusidae): A New Crab Intermediate Host of Paragonimus mexicanus in Manabí Province, Ecuador

To determine that Paragonimus sp. is actively transmitted in a tropical area of the Pacific region of Ecuador where human cases of pulmonary paragonimiasis have recently been documented, a total of 75 freshwater crabs were collected from 2 different streams in the Pedernales area of Manabí Province, Ecuador. All collected crabs were identified as Hypolobocera guayaquilensis based on morphological characteristics of the male gonopods. The hepatopancreas of each crab was examined by compressing it between 2 glass plates followed by observation under a stereomicroscope. Excysted Paragonimus metacercariae were detected in 39 (52.0%) crabs and their densities varied from 1 to 32 per infected crab. There was a positive relationship between crab size and metacercarial density. Sequences of the second internal transcribed spacer region of the ribosomal RNA gene of the Paragonimus metacercariae obtained in this study were identical to those of Paragonimus mexicanus deposited in the DDBJ/EMBL/GenBank database. Thus, the present study is the first to confirm that the crab species H. guayaquilensis is the second intermediate host of P. mexicanus in Manabí Province, Ecuador. Because this crab might be the possible source of human infections in this area, residents should pay attention to improper crab-eating habits related with a neglected parasitic disease, i.e., paragonimiasis.