Importance of database curation in taxonomic assignation of 16S data.
Microbial identification is the key component to microbial community analysis. Since mid-2000s with the advent of Next-generation sequencing techniques it has been necessary to use increasingly refined and complete databases to uniquely assign the taxonomy of each sequence or taxonomic unit. In this study we evaluate the relevance of the database curation in this assignation process.
Prosthetic valve endocarditis due to Candida parapsilosis- a rare case report
Editorial office note: The received date on the following peer review reports Reviewer 1 (https://doi.org/10.1099/acmi.0.000462.v2.3) and Reviewer 3 (https://doi.org/10.1099/acmi.0.000462.v2.1) have been amended to display correctly in the Review Status Breakdown box. These reports were received on 12 October 2022 and 27 September 2022 respectively.
Fungal endocarditis is a rare and fatal condition most frequently caused by the Candida and Aspergillus species. Fever and changing heart murmur are the most common clinical manifestations. The diagnosis of fungal endocarditis is challenging especially that of prosthetic valve endocarditis is extremely difficult. The optimal management of the condition still remains debatable. We hereby present a case of prosthetic valve endocarditis caused by Candida parapsilosis managed empirically with liposomal amphotericin B and later on was shifted to combination therapy with high dose echinocandin and fluconazole but had a fatal outcome because he could not undergo timely surgical intervention. Treating Candida parapsilosis endocarditis cases are difficult because of their ability of biofilm production on native and prosthetic heart valves. A combined approach consisting of high index of clinical suspicion early diagnosis by using serological markers followed by culture or PCR and prompt initiation of appropriate antifungals may aid in improving the outcome.
Prevalence of Hepatitis B Virus Surface Antigen (HBsAg) among the Out-patients in a Tertiary Institution in Nigeria.
Hepatitis B virus (HBV) infection is considered to be the ninth leading cause of death and is one of the major public health problems in the world. The prevalence of hepatitis in Nigeria has been rated to be 8.3%. Hence this study investigate the prevalence of hepatitis B virus infection in a tertiary institution in Oyo State. Informed consent were obtained from the out-patients attending institution clinics. The design was a cross-sectional study and questionnaires were administered to investigate their knowledge and attitude towards safer practices against the spread of HBV. The seroprevalence of the participants were examined with LabAcon rapid kit with 98.2% - 100% and 97.2% - 99.8 for specificity and sensitivity respectively. The population distribution was observed to 45 (90%) female and 5 (10%) male. The prevalence of the HBsAg was found to be 10%. The prevalence of HBsAg among female participants was 11.1%. The relationship between knowledge of HBV infection and sex was insignificant (0.546) however the relationship between knowledge of HBV infection and age was significant (0.004). The occurrence of HBV infection is still area of concern because of the possibility of the virus to spread. This spread could better be curtailed based on regular awareness about the infection.
Building Blocks of Biofilms – an engaging and hands-on microbiology outreach activity for school children and the general public
Biofilms are naturally occurring communities of microorganisms attached to a surface and often embedded in a matrix of self-produced polymeric substances. Biofilms are widely implicated in human infections particularly on prostheses and medical implants. Such biofilms are difficult to eradicate often leading to replacement of the prosthesis and resulting in a significant burden to healthcare. Here we present a fun and engaging interactive activity targeted toward primary school/early secondary school children introducing the concept of natural and healthcare-associated biofilms using dental plaque as an archetypal example. Dental plaque forms as a result of poor oral/dental hygiene and develops according to a typical series of defined stages: attachment and adherence to the surface followed by colonisation and maturation of the biofilm structure and eventually dispersal. This activity uses dental disclosing tablets to visualise real biofilms (plaque) on the participants teeth and uses interlocking building-blocks to represent microorganisms where children build three-dimensional ‘biofilms’ of varying shapes and structural integrities. Each of the stages of development are discussed in detail and after building the biofilms balls of different shapes sizes and weights can be used as ‘antimicrobials’ to disrupt the biofilm structure. The outcomes of the activity are to enhance knowledge and general understanding of biofilms; their ubiquitous presence in the natural environment development implications in healthcare and challenges of treatment. The various ‘antimicrobial’ balls also provide a basis to introduce and discuss drug selection for infections and the importance of using the correct antimicrobial for different infections to avoid development of resistance.
Detection of OXA-48 and NDM-1 genes in ESBL producing Ochrobactrum anthropi from urine samples of Out-patients.
Background: Treatment of the urinary tract infections (UTIs) is vast becoming worrisome because prominent antibiotic resistance among the bacteria involve in the infection. Species of Ochrobacterum are also involve in the infection affecting urinary tract and profusely resistance to antibiotics.
Methods: Urine samples were collected from out-patients of some hospitals and the bacteria were isolated and identified based on the morphological and biochemical characteristics. The most resistant isolate was selected for molecular identification by amplifying 16S rRNA gene. The bacterial isolates were subjected to antibiotic susceptibility assay and interpreted using standard of CLSI. The resistant isolate was evaluated for the presence of ESBL genes MBL genes biofilm gene and efflux pump gene.
Results: Fourteen (14) Ochrobactrum spp. were isolated from urine samples. Only 3 (21%) of the isolates were resistance to all the twenty-one (21) antibiotics while 7 (50%) were resistance to twenty (20) antibiotics. However resistance was observed to atleastnine (9) antibiotics in all the antibiotics by the bacterial isolates. The selected isolated was identified to be Ochrobactrum anthropic strain U0145 with occurrence of blaTEMblaSHV and blaCTX-M-9 as ESBLs genes while blaOXA-48 and blaNDM-1 genes as MBLs genes however there was no occurrence of GES 1-911 gene. The bacterium possessed biofilm production gene while there was no efflux pump gene.
Conclusion: O. anthropi is becoming prominent among the bacteria responsible for nosocomial-associated infection especially when catheter is involved and treatment may be tasking therefore adequate infection control should be put in place.
Evaluation of five commercial DNA extraction kits using Salmonella as a model for implementation of rapid Nanopore sequencing in routine diagnostic laboratories
Oxford Nanopore long-read sequencing offers advantages over Illumina short-reads for the identification and characterization of bacterial pathogens for outbreak detection and surveillance activities within a diagnostic public health laboratory context. Compared to Illumina Nanopore is more cost-effective for small batches has a lower capital cost and has a faster turnaround time in addition to the ability to assemble complete bacterial genomes. The quantity and quality of DNA required for Nanopore sequencing is higher than Illumina and DNA extraction methods recommended for obtaining high molecular weight DNA are different from those typically used in diagnostic laboratories. Using a Salmonella isolate with a previously closed PacBio genome as a model Enterobacteriaceae organism we evaluated the quantity quality and fragmentation of five commercial DNA extraction kits. Nanopore sequencing performance was evaluated for the top three methods: Qiagen EZ1 DNA Tissue Qiagen DNeasy Blood and Tissue and a modified in-house version of the MasterPure Complete DNA and RNA purification. To evaluate the effect of post-extraction DNA purification methods we subjected extracted DNA from the three selected extraction methods to purification by AMPure beads or ethanol precipitation and compared these outputs with untreated DNA as a control. All methods are suitable for routine Whole Genome Sequencing (WGS) since all 60 replicates had very high genome recovery rates with ≥ 98% of the reference genome covered by mapped Nanopore reads. For 85% of the replicates assembly was able to produce a complete circular chromosome using either Flye or Canu. In most cases it is recommended to move directly from extraction to sequencing as untreated DNA had the highest rates of genome closure regardless of extraction method. Using our evaluation criteria the Qiagen DNeasy Blood and Tissue kit was found to be the best overall method due to its low-cost ability to scale from single tube to 96-well plates and high consistency in yield and sequencing performance.
Vaccine-induced neutralizing antibodies against SARS-CoV-2 Omicron variant isolated in Osaka, Japan
To study vaccine-induced neutralizing antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants isolated in Osaka Japan microneutralization tests were performed on serum samples from 32 subjects who received a second dose of vaccination and 10 of those who received the third dose of vaccination. Geometric Mean Titres (GMTs) for the D614G strain Alpha variant Delta variant and Omicron BA.1 of the subjects after the second dose of vaccination were 19.5 21.8 6.3 and 2.0 respectively. The GMT for the Delta variant was significantly lower than that for the D614G strain and Alpha variant and the GMT for the Omicron BA.1 was significantly lower than that for the Delta variant. Among the subjects who received three doses of vaccination the GMTs for the Omicron BA.1 (62.8) and BA.2 (38.6) were significantly higher than that for the Omicron BA.1 after the second dose. Thus in the present study the second dose of vaccination induced neutralizing antibodies against SARS-CoV-2 strains and the reactivity of neutralizing antibodies to the variants was thought to be enhanced by the third dose of vaccination. The serum samples used in this study will be useful in evaluating the reactivity of vaccine-induced antibodies to newly emerging variants.
Gene Sequencing: Key for Analyzing Mutation in SARS-CoV-2
Genomic sequencing is a sole platform that recognizes and characterizes novel mutants of microorganisms existing in various clinical samples. The main objective of this article is to summarize and provide relevant information regarding genomic sequencing platforms that are widely applied for analyzing mutations in SARS-CoV-2. Various data and literature were reviewed from scientific database like PubMed Scopus Google scholars and internet in order to provide collective and reliable information on genomic sequencing. Various Variants of Concern (VOC) and Variants of Interest (VOI) with higher transmissibility and infectivity are detected around the globe among which Nepal is also at high risk stage. According to the Ministry of Health and Population (MOHP) delta variants (B.1.617.2) alpha variants (B.1. 1.7) kappa variants(B.1.617.1) and newly emerged sub lineage of delta variants (A.Y.1) are the predominant one that are circulating in the country. Therefore severe acute respiratory syndrome coronavirus 2 whole genome sequencing Next generation sequencing (Illumina MiSeq. and Oxford Nanopore Minion) or Sanger method based on amplicon sequencing are the best optional platforms for the detection of VOC and VOI and to continue monitoring of widespread SARS-CoV-2 for proper therapy and prevention of COVID-19. Besides variations occurring in the fragment of virus genome may produce gene dropout obstructing with laboratory molecular assays and epidemiological surveys therefore it is very crucial to detect those markers with the aid of gene sequencing methods in order to establish higher molecular testing strategies.
INTERMEDIATE VAGINAL FLORA IS ASSOCIATED WITH HIGHLY ACTIVE ANTIRETROVIRAL THERAPY IN HIV POSITIVE PREGNANT WOMEN ON HIGHLY ACTIVE ANTIRETROVIRAL THERAPY IN KISUMU, KENYA.
Introduction A number of studies have linked Highly Active Antiretroviral Therapy (HAART) to adverse pregnancy outcome but the mechanism is not well understood. Hypothesis This study postulated abnormal vaginal flora (AVF) as the link between HAART and adverse pregnancy outcome. Objective The study was designed to determine correlates of AVF with special emphasis on the association between AVF and HAART in HIV-1 positive pregnant women on HAART in Kisumu Kenya. Method This cross sectional study was carried out at Lumumba sub county hospital. The facility receive patients from Kisumu county and its surrounding; a region with high HIV and BV prevalence plus high maternal and neonatal morbidities and mortalities. Willing 76 pregnant women who started HAART on or before first trimester were enrolled after signing informed consent forms. Using formatted questionnaire participants were interviewed on demographic clinical and behavioral information. The samples were collected in second trimester and BV determined by Nugent’s criteria. Fisher Exact Chi square test and multinomial logistic regression was used to determine the association of HAART and participants characteristics with AVF. Results Vaginal douching any time earlier than two weeks to specimen collection [9.67(1.03-90.41)] and non efavirenz based HAART [24.57(2.39-253.14)] were positively associated with BV and intermediate vaginal flora respectively while self-employment [0.06(0.01-0.88)] and one sexual partner in the last two months [0.13(0.02-0.70)] being negatively associated with intermediate vaginal flora. Meanwhile non efavirenz based HAART [17.10(1.18-247.34)] and vaginal douching anytime earlier than two weeks to specimen collection [12.70(1.10-146.)] had independent positive association with intermediate vaginal flora and BV respectively. Conclusion This study found a positive independent association between HAART and intermediate vaginal flora. Similar to BV it has been discovered that intermediate vaginal flora is linked to poor pregnancy outcome. This study has therefore proved that HAART alters vaginal flora leading to adverse pregnancy outcome.
Comparison of diagnostic accuracy of Xpert MTB/RIF and Geno type MTBDRplus line probe assayfor diagnosis of tuberculous pleural effusion
Background: Tuberculous pleural effusion (TPE) occurs in up to 25% of TB patients. Owing to the pauci-bacillary nature of the pleural fluid the diagnosis of TPE is a challenge. Newer diagnostic tools are required for the rapid diagnosis of TPE.
Objectives: To compare the sensitivity and specificity of Xpert MTB/RIF and Geno type MTBDRplus line probe assay (MTBDRplus) for diagnosing TPE.
Methods: A prospective cross-sectional study was performed at Aga Khan University Hospital Karachi Pakistan from August 2014 to January 2016. Patients with suspected TPE were recruited on the basis of history exudative lymphocytic nature of effusion and raised adenosine daminase level. Pleural fluid samples were tested for AFB smear culture Xpert MTB/RIFand MTBDR plus .
Results: We enrolled 99 patients with mean age of 50.4±20.3 years. AFB culture was positive in 14 (14.14%) cases. Considering AFB culture as Gold standard the sensitivity of Xpert MTB/RIF was found to be 57.14% (95% CI: 28.86 – 82.34%) and specificity was 97.65% (95% CI: 91.76 - 99.71%) and the sensitivity of MTBDR plus was 35.71% (95% CI: 12.76 – 64.86%) and specificity was 98.82% (95% CI: 93.62 - 99.90%).The sensitivity of Xpert MTB/RIF in TPE was higher than MTBDRplus (p 0.013) while specificity was similar.
Conclusion: Xpert MTB/RIF is more sensitive for detecting TPE than MTBDRplus and AFB smear microscopy. A multicenter large-sample study is needed to evaluate this method for early TPE diagnosis.
Bacillus circulans septicemia in a premature infant: a case report
In immunocompromised patients Bacillus circulans is primarily regarded an opportunistic pathogen. This is a case report of septicaemia caused by Bacillus circulans in an extremely low birth weight preterm baby with respiratory distress. Although a laboratory contaminant but in immunocompromised host isolation of B. circulans should not be overlooked.
Deep phylotaxonogenomic revision of the genus Xanthomonas within family Xanthomonadaceae and proposal of novel family Frateuriaceae within order Xanthomonadales
Genus Xanthomonas is primarily comprise phytopathogenic species. In a recent study by carrying out deep phyto-taxonogenomics we reported that even the genera Xylella Stenotrophomonas and Pseudoxanthomonas are miss-classified and belong to genus Xanthomonas. Hence to understand the breadth of the genus we carried out deep phylo-taxonogenomics of the order Xanthomonadales. Such investigation revealed that at least four more genera belong to genus Xanthomonas with prominent being Lysobacter. Further order level deep phylo-taxonogenomics revealed two major families. One being the original family Xanthomonadaceae and other is proposed as Frateuriaceae fam. nov. as synonym of family Rhodanobacteraceae with novel genus Frateuria gen. nov.
Human brucellosis: An observational study from a tertiary care centre in northern India
Brucellosis is a neglected zoonotic disease that affects both humans and animals. In humans brucellosis begins with chronic illness leading to great financial losses from not being able to work well and continued treatment costs. Joint pain is the common presentation of brucellosis and there are several risk factors associated with brucellosis. But few such researches have come from northern India. We studied to detect and characterize the Brucella species from patients having complaints of joint pain and also to know the potential causes of human brucellosis. A total of 200 blood samples were collected from the participants having joints pain from September 2019 to September 2021 at Gandhi memorial and associated hospital of King George’s Medical University Lucknow Uttar Pradesh and tested by serology for anti brucella IgM and IgG molecular tests by RTPCR conventional PCR and automated blood culture system. The study showed positive results by anti-brucella IgM ELISA 19 (9.5%) anti brucella IgG 23 (11.5%) and of those 1 (0.5%) was positive for both anti-brucella IgM and anti-brucella IgG ELISA. Out of 19 anti-brucella IgM ELISA positive 8 (4%) samples were positive for PCR/ RT-PCR and that was negative for anti-brucella IgG ELISA. All blood culture reports of all patients were negative. Our study concluded that anti-brucella IgM ELISA was more accurate than anti-brucella IgG ELISA in detecting human brucellosis. Consumption of animal products (i.e. milk a dairy product of cow buffalo goat and meat of goat) and contact with animals were the main risk factors that were identified for brucella disease.
Cloning and expression of α-Amylase from novel specie, Enterobacter xiangfangensis; locally isolated from Pakistani flora
Living in an era of industrialization there is an utmost need to replace chemical catalysts with biocatalysts owing their cost effectiveness milder reaction conditions eco-friendly nature and specificity. α-amylase is one of the top commercial enzymes of industrial importance. It hydrolyzes internal α (1 4) glycosidic linkages in polysaccharides and yields oligosaccharides of varying length. The aim of present study was to clone and express α-amylase from a novel species Enterobacter xiangfangensis. Amplified product was cloned by A and T overhangs and transformed into cloning host; E. coli DH5α cells. Gene encoding α-amylase (1488bp) was sequenced to confirm the adequate amplification digested with BamH1 and Xho1 cloned into expression vector; pET28b (+) and transformed into expression host; E. coli strain BL21 CodonPlus (DE3) cells.
The expression of α-amylase was induced by IPTG and optimized with varying IPTG concentrations and its induction intervals. The maximum expression was observed after 8 hours of 1mM IPTG induction. The results showed that α-amylase from novel specie Enterobacter xiangfangensis has a potential to expresses itself in heterologous expression system so this recombinant α-amylase is ray of hope for enormous downstream applications.
The emerging human monkeypox in Thailand: a mini-review
Monkeypox virus infection is a newly emerging viral infectious disease that has become the current public health threat. Formerly monkeypox was a crucial zoonotic virus endemic limited to the central and west African regions never been reported in Southeast Asia. Humans can be infected with the monkeypox virus by contact with infected humans and animals. However the recent emergence of monkeypox worldwide including in Thailand is of significant interest and shall be surveillance. In addition the recent evidence from the recent outbreaks worldwide demonstrates that infection can be transmitted by sexuality. The author reviewed and represented important information on this disease in this mini-review.
Whole Genome Sequencing Based Characterization of Streptomyces sp. 6(4): focus on natural products
We have sequenced the whole genome of Streptomyces sp. 6(4) isolated from tomato roots that presents antifungal activity against phytopathogenic fungi mainly against Bipolaris sorokiniana . The genome has almost 7 Mb and 3368 hypothetical proteins that were analyzed and characterized in Uniprot with emphasize in biological compounds. MLST analysis were performed in effort to characterize and identify this isolate resulting in a new ST classified as ST64. Phenetic and phylogenetic trees were constructed to investigate Streptomyces sp. 6(4) evolution and sequence similarity and the isolate is a strain closer to Streptomyces prasinus and Streptomyces viridosporus . It is known that Streptomyces genera comprises a huge metabolism capacity with the presence of cryptic genes. These genes are usually present in clusters who are responsible for production of a high diversity of natural products mainly antibiotics. In addition 6(4) showed 11 biosynthetic gene cluster through antiSMASH including 3 clusters PKS and NRPS type.
Virome diversity in rodents and their zoonotic viral infectious diseases: an essential reservoir of emerging and re-emerging viruses infecting humans
Rodents are the largest mammalian order (Rodentia) and are extraordinarily diverse. Several rodent species are closely relevant to humans such as destroying agricultural products and spreading viral pathogens. Rodents are essential reservoirs for zoonotic viruses with notable instances including the Lassa virus and hantaviruses. A few characteristics of rodents such as the number of species colonial population communities and immune response make them an exquisitely suitable essential reservoir of viruses. Mainly infecting humans reflects direct contact with wildlife including secretion and body fluids. Here the author summarises the current understanding of the virome diversity in rodents and their zoonotic viral infectious diseases. However a significant consideration is that although rodents maintain multiple viruses some rodent viruses have not caused human disease.
Shewanella putrefaciens: a rare cause of purulent otorrhea !
Shewanella putrefaciens is a Gram-negative non-fermenting motile and oxidase-positive bacillus. Its incrimination in human pathology is very rare with a resurgence of Shewanella infections in recent years.We report the first case in Morocco of a purulent otorrhea caused by Shewanella putrefaciens rebel to conventional treatment occurring in a 25-year-old female afebrile without deterioration of the general state and with a notion of sea bathing. We also describe the bacteriological characteristics and antibiotic susceptibility results of the isolate.
Genomic analysis of the progenitor strains of Staphylococcus aureus RN6390
RN6390 is a commonly used laboratory strain of Staphylococcus aureus derived from NCTC8325. In this study we sequenced the RN6390 genome and compared it to available genome sequences for NCTC8325. We confirmed that three prophages Φ11 Φ12 and Φ13 which are present in NCTC8325 are absent from the genome of RN6390 consistent with the successive curing events leading to the generation of this strain. However we noted that a separate prophage is present in RN6390 that is not found in NCTC8325. Two separate genome sequences have been deposited for the parental strain NCTC8325. Analysis revealed several differences between these sequences in particular between the copy number of esaG genes which encode immunity proteins to the type VII secreted anti-bacterial toxin EsaD. Single nucleotide polymorphisms were also detected in ribosomal RNA genes and genes encoding microbial surface components recognizing adhesive matrix molecules (MSCRAMM) between the two NCTC8325 sequences. Comparing each NCTC8325 sequence to other strains in the RN6390 lineage confirmed that sequence assembly errors in the earlier NCTC8325 sequence are the most likely explanation for most of the differences observed.
Methicillin-Resistant Staphylococcus aureus Carriage on The Hands of Healthcare Workers: An Assessment for Hand Hygiene Practices in Jos, Nigeria.
Introduction Methicillin-resistant Staphylococcus aureus (MRSA) is capable of causing a wide range of infections. Contaminated hands of healthcare workers (HCWs) act as a potential source of MRSA transmission in hospitals. Aim This study aims at detecting the carriage of MRSA on the hands of HCWs during patient care and the evaluation of the effective practice of hand hygiene protocol. Methodology This study was a cross-sectional point (prevalence) study done in wards and intensive care units (ICUs) of a tertiary care hospital. Cultures from hand swabs were obtained from HCWs during clinical rounds without prior information about the procedure. A second culture was also taken after the use of an alcohol-based hand rub or handwash (after 1 minute of use). Result Of a total of 145 HCWs screened 24 (17%) were positive for MRSA. Doctors were 9 while nurses were 10. Six (4%) HCWs remained positive for MRSA after washing their hands or using an alcohol-based hand rub. Conclusion Commitment to preventing healthcare-associated infections requires that regular monitoring of hand hygiene practices be in place. A rapid diagnostic protocol such as MRSA chromogenic agar has a shorter turnaround time compared to standard laboratory workflow in identifying MRSA. Evaluation of the transmission rate of MRSA from HCWs to patients will be encouraged in further studies.