A2 Isolates Research Articles

Isolasi dan Karakterisasi Plant Growth Promoting Rhizobacteria dari Perakaran Kelapa Sawit pada Lahan Gambut

Peatlands have characteristics of low pH and lack nutrients. Oil palm is the main plant commodity in peatland management. Oil palm roots have been known to be a nutrition source for the growth of soil microbes, especially bacteria around their roots or PGPR. PGPR are a group of bacteria that play an important role in supporting plant growth and health. The purpose of this research was to obtain PGPR potential from oil palm roots which can be used as candidates for biofertilizer agents. In this study, the isolation and selection of PGPR isolate from oil palm roots on oil palm plantations in Central Kalimantan were carried out based on their plant growth-promoting traits, including the activity of producing Indole Acetic Acid (IAA), phosphate solubilizing, N-fixing, K solubilizing, siderophore production, ACC deaminase activity, proteolytic activity, cellulolytic activity, and ligninolytic activity. A total of 17 isolates were selected to be tested for their multiple activities ability. The final results of the PGPR characterization showed that of the seventeen isolates, all isolates had PGPR activity at least three different abilities. From the seventeen isolates, it was found that the SW 5.5 PK 3A isolate had the highest IAA production activity (58,50 ppm), SW 4.10 PK 1A isolate had the highest K solubilizing index (3,16), SW 8.5 PK 1A isolate had both the highest P solubilizing index (3,73) and the highest siderophore zone index (5,20), SW 4.11 PK isolate had the highest proteolytic index (4,80), and SW 4.10 PK 1A.P isolates had the highest cellulolytic index (5,11).

Characterization and fungicides sensitivity of Phytophthora cinnamomi isolates causing avocado root rot in Zitácuaro, Michoacán

<em>Phytophthora cinnamomi</em> is the pathogen most frequently associated with avocado root rot. In Zitácuaro, Michoacán, production has increased by 19.8%; however, there are no studies of root rot in this area. The objective of the study was to characterize the isolates obtained from avocado roots and assess the sensitivity to fungicides. Samples from 5 avocado orchards were collected, sampling 5 trees per orchard (a total of 25 samples). The samples isolated were characterized morphological and molecularly. Mating type was analyzed using reference isolates of<em> P. cinnamomi</em> A1 (isolate from camelia) and A2 (isolate from avocado). To confirm the pathogenicity, tests were performed on avocado fruits with the isolates. The sensitivity of 15 isolates to potassium phosphite and to metalaxyl-M at different concentrations was evaluated<em> in vitro</em>. In a subgroup of six isolates, it was evaluated whether there was a relationship between growth rate and potassium phosphite sensitivity. Fifteen isolates were obtained with coenocytic coraloid mycelium, chlamydospores, sporangia without papilla, ovoid to ellipsoid, with internal proliferation, heterothallic with mating type A2, with amphigynous antheridia and plerotic oospores, characteristics consistent with <em>P. cinnamomi</em>. The inoculated isolates were pathogenic on avocado fruits. The isolates were more sensitive to potassium phosphite than to metalaxyl-M, with mean EC50 values of 24.62 and 0.215 ?g mL-1 of i.a., respectively. No relationship was observed between growth rate and potassium phosphite sensitivity. It is necessary to obtain a greater number of<em> P. cinnamomi</em> isolates for virulence studies.

Isolation and characterization of nitrogen-fixing bacteria from soil sample in Dhaka, Bangladesh

Biological nitrogen (N2) fixation is very essential for limiting the growth of plants and agricultural crops. The present study was conducted to potentially isolate N2-fixing bacteria from garden soil sample at Stamford University Bangladesh, Siddeswari, Dhaka. Here, we used culture-dependent method to perform this experiment. Firstly, we collected garden soil sample, diluted and inoculated in N2-free Jensen’s media by maintaining the aseptic procedure. We obtained 5 different colonies from soil samples. We cultured the isolates in N2-free Jensen’s media containing bromothymol blue (BMB) and also, in Yeast Extract Mannitol Agar (YEMA) media containing congo red to confirm nitrogen fixation capacity. We collected the colony characteristics of all the isolates. Only 1A isolate showed good growth after 24 h of incubation among all the isolates. We performed ammonification test with Nessler reagent to confirm N2-fixing ability for our selected isolates. The 1A isolate was positive in ammonification test. Culture, microscopy and biochemical tests were performed to identify isolate 1A. This isolate was presumptively identified as Azotobacter sp. In the present study, Azotobacter sp. that was isolated from the soil sample was found to be a potential N2-fixing bacterium. Isolate 1A can be used for N2-fixation to boost production of crops. Stamford Journal of Microbiology, Vol.11 (1) 2021: 11-13

Isolation, partial purification and characterization of phospholipase A2 from Naja Katiensis venom

The most effective and acceptable therapy for snakebite victims is the immediate administration of antivenin which is limited by problems of hypersensitivity reactions in some individuals and its inability to resolve the local effects of the venom. The aim of this study was to isolate, partially purify and characterize phospholipase A2 from Naja Katiensis venom. Phospholipase A2 was partially purified via a two-step process: gel filtration on Sephadex G-75 and ion exchange chromatography using CM Sephadex, and subjected to SDS-PAGE analysis. From the results, the specific activity of the partially purified PLA2 decreased from 0.67μmol/min/mg in crude venom to 0.29μmol/min/mg after ion exchange chromatography with a yield of 5% and purification fold of 0.43. The optimum temperature of the purified PLA2 was found to be 35ºC and optimum p.H of 7. velocity studies for the determination of kinetic constants using L-a-lecithin as substrate revealed a Km of 1.47mg/ml and Vmax of 3.32μ moles/min/mg. The sodium dodecyl sulphate polyacrylamide gel electrophoresis of the purified PLA2 showed a distinct band with molecular weight estimated to be 14KDa. In conclusion, the present study shows that phospholipase A2 was isolated, purified and characterized. This may serve as a promising candidate for future development of a novel anti-venin drug.

Relationship Between Serotypes and Antimicrobial Nonsusceptibility of Streptococcus pneumoniae Clinical Isolates in Tunisia.

Streptococcus pneumoniae remains a significant cause of morbidity and mortality worldwide despite the overall success of the vaccine programs. In Tunisia, pneumococcal conjugate vaccines (PCV)10 was introduced in the national immunization program in April 2019. We sought to determine the relationship between serotypes and antimicrobial nonsusceptibility of S. pneumoniae isolates recovered from clinical samples in the prevaccination period in the south of Tunisia. A total of 504 nonduplicate S. pneumoniae isolates collected between 2012 and 2018 were tested for antimicrobial susceptibility, among them 439 (87.1%) were serotyped. The most common serotypes were 19F (17.8%), 14 (15.3%), 3 (9.1%), 19A (8.2%), and 23F (7.3%). The proportions of isolates with serotypes covered by PCV7, PCV10, and PCV13 were 55.4%, 56.3%, and 77.9%, respectively. Three-quarters (74.4%) of pneumococcal isolates were nonsusceptible to penicillin, and about half (54.8%) were multidrug resistant. Penicillin nonsusceptibility was observed for all 19A and 23F isolates, and was significantly associated with serotypes 19F (odds ratio [OR]: 33.7) and 14 (OR: 8.7). A significant association with multidrug resistance was noted for serotypes 19A (OR: 10), 19F (OR: 9.4), 23F (OR: 8.6), and 6B (OR: 5.2). The alarming rates of pneumococcal antimicrobial nonsusceptibility and the strong association with the most prevalent serotypes compel microbiologists to monitor the impact of the PCV10 introduced recently in our national immunization program.

First Report of Stem and Foliage Blight of Chrysanthemum Caused by Phytophthora drechsleri in the United States.

Chrysanthemum (Chrysanthemum × morifolium) plants exhibiting stem and foliage blight were observed in a commercial nursery in eastern Oklahoma in June 2019. Disease symptoms were observed on ~10% of plants during a period of frequent rain and high temperatures (26-36°C). Dark brown lesions girdled the stems of symptomatic plants and leaves were wilted and necrotic. The crown and roots were asymptomatic and not discolored. A species of Phytophthora was consistently isolated from the stems of diseased plants on selective V8 agar (Lamour and Hausbeck 2000). The Phytophthora sp. produced ellipsoid to obpyriform sporangia that were non-papillate and persistent on V8 agar plugs submerged in distilled water for 8 h. Sporangia formed on long sporangiophores and measured 50.5 (45-60) × 29.8 (25-35) µm. Oospores and chlamydospores were not formed by individual isolates. Mycelium growth was present at 35°C. Isolates were tentatively identified as P. drechsleri using morphological characteristics and growth at 35°C (Erwin and Ribeiro 1996). DNA was extracted from mycelium of four isolates, and the internal transcribed spacer (ITS) region was amplified using universal primers ITS 4 and ITS 6. The PCR product was sequenced and a BLASTn search showed 100% sequence similarity to P. drechsleri (GenBank Accession Nos. KJ755118 and GU111625), a common species of Phytophthora that has been observed on ornamental and vegetable crops in the U.S. (Erwin and Ribeiro 1996). The gene sequences for each isolate were deposited in GenBank (accession Nos. MW315961, MW315962, MW315963, and MW315964). These four isolates were paired with known A1 and A2 isolates on super clarified V8 agar (Jeffers 2015), and all four were mating type A1. They also were sensitive to the fungicide mefenoxam at 100 ppm (Olson et al. 2013). To confirm pathogenicity, 4-week-old 'Brandi Burgundy' chrysanthemum plants were grown in 10-cm pots containing a peat potting medium. Plants (n = 7) were atomized with 1 ml of zoospore suspension containing 5 × 103 zoospores of each isolate. Control plants received sterile water. Plants were maintained at 100% RH for 24 h and then placed in a protected shade-structure where temperatures ranged from 19-32°C. All plants displayed symptoms of stem and foliage blight in 2-3 days. Symptoms that developed on infected plants were similar to those observed in the nursery. Several inoculated plants died, but stem blight, dieback, and foliar wilt were primarily observed. Disease severity averaged 50-60% on inoculated plants 15 days after inoculation. Control plants did not develop symptoms. The pathogen was consistently isolated from stems of symptomatic plants and verified as P. drechsleri based on morphology. The pathogenicity test was repeated with similar results. P. drechsleri has a broad host range (Erwin and Ribeiro 1996; Farr et al. 2021), including green beans (Phaseolus vulgaris), which are susceptible to seedling blight and pod rot in eastern Oklahoma. Previously, P. drechsleri has been reported on chrysanthemums in Argentina (Frezzi 1950), Pennsylvania (Molnar et al. 2020), and South Carolina (Camacho 2009). Chrysanthemums are widely grown in nurseries in the Midwest and other regions of the USA for local and national markets. This is the first report of P. drechsleri causing stem and foliage blight on chrysanthemum species in the United States. Identifying sources of primary inoculum may be necessary to limit economic loss from P. drechsleri.

Late Blight Resistance Evaluation and Genome-Wide Assessment of Genetic Diversity in Wild and Cultivated Potato Species

Late blight, caused by the oomycete Phytophthora infestans, is the most devastating disease in potato-producing regions of the world. Cultivation of resistant varieties is the most effective and environmentally friendly way to control potato late blight disease, and identification of germplasms with late blight resistance and clarification their genetic relationship would promote the development of the resistant varieties. In this study, a diverse population of 189 genotypes with potential late blight resistance, consisting of 20 wild species and cultivated Solanum tuberosum Andigenum group and Chilotanum group, was screened for the presence of late blight resistance by performing challenge inoculation with four Phytophthora infestans isolates including one 13_A2 isolate, CN152. Ten elite resources with broad-spectrum resistance and 127 with isolate-specific resistance against P. infestans were identified. To improve the available gene pool for future potato breeding programs, the population was genotyped using 30 simple sequence repeat (SSR) markers covering the entire potato genome. A total of 173 alleles were detected with an average of 5.77 alleles per locus. Structure analysis discriminated the 189 potato genotypes into five populations based on taxonomic classification and genetic origin with some deviations. There was no obvious clustering by country of origin, ploidy level, EBN (endosperm balance number) value, or nuclear clade. Analysis of molecular variance showed 10.08% genetic variation existed among populations. The genetic differentiation (Fst) ranged from 0.0937 to 0.1764, and the nucleotide diversity (π) was 0.2269 across populations with the range from 0.1942 to 0.2489. Further genotyping of 20K SNP array confirmed the classification of SSRs and could uncover the genetic relationships of Solanum germplasms. Our results indicate that there exits abundant genetic variation in wild and cultivated potato germplasms, while the cultivated S. tuberosum Chilotanum group has lower genetic diversity. The phenotypic and genetic information obtained in this study provide a useful guide for hybrid combination and resistance introgression from wild gene pool into cultivated species for cultivar improvement, as well as for germplasm conservation efforts and resistance gene mining.

Functional characterization of a gene cluster responsible for inositol catabolism associated with hospital-adapted isolates of Enterococcus faecium.

Enterococcus faecium is a nosocomial, multidrug-resistant pathogen. Whole genome sequence studies revealed that hospital-associated E. faecium isolates are clustered in a separate clade A1. Here, we investigated the distribution, integration site and function of a putative iol gene cluster that encodes for myo-inositol (MI) catabolism. This iol gene cluster was found as part of an ~20 kbp genetic element (iol element), integrated in ICEEfm1 close to its integrase gene in E. faecium isolate E1679. Among 1644 E. faecium isolates, ICEEfm1 was found in 789/1227 (64.3 %) clade A1 and 3/417 (0.7 %) non-clade A1 isolates. The iol element was present at a similar integration site in 180/792 (22.7 %) ICEEfm1-containing isolates. Examination of the phylogenetic tree revealed genetically closely related isolates that differed in presence/absence of ICEEfm1 and/or iol element, suggesting either independent acquisition or loss of both elements. E. faecium iol gene cluster containing isolates E1679 and E1504 were able to grow in minimal medium with only myo-inositol as carbon source, while the iolD-deficient mutant in E1504 (E1504∆iolD) lost this ability and an iol gene cluster negative recipient strain gained this ability after acquisition of ICEEfm1 by conjugation from donor strain E1679. Gene expression profiling revealed that the iol gene cluster is only expressed in the absence of other carbon sources. In an intestinal colonization mouse model the colonization ability of E1504∆iolD mutant was not affected relative to the wild-type E1504 strain. In conclusion, we describe and functionally characterise a gene cluster involved in MI catabolism that is associated with the ICEEfm1 island in hospital-associated E. faecium isolates. We were unable to show that this gene cluster provides a competitive advantage during gut colonisation in a mouse model. Therefore, to what extent this gene cluster contributes to the spread and ecological specialisation of ICEEfm1-carrying hospital-associated isolates remains to be investigated.

Phylogenetic tree of NS5A gene of hepatitis C virus from infected Iraqi patients

Hepatitis C is a liver illness caused by the hepatitis C virus (HCV). Acute and chronic hepatitis can be caused by the virus. Hepatitis C infection affects an estimated 71 million people worldwide. Two hundred blood samples were collected (128 males and 72 females), ranged in age from 4 to 80 years and diagnosed through a rapid test and the real time-PCR from the teaching medical city hospital in Baghdad from October 2018 until April 2019. The results showed that 89 samples were positive for HCV by serological tests, and other samples are not. Also, the most frequent age of infection of HCV was between 51 and 80 years. In all samples, the partial region of the NS5A gene was successfully amplified and sequenced. PCR-sequencing revealed three subtyping were; subtype 1a, 1b, and 4a. Phylogenetic analysis revealed subtype concordance of 99 % with a bootstrap hepatitis C virus subtype 1b isolate AASN1 Iraq (ID:MT554101.1). The clusters including HCV subtype 1a isolate AASN20 Iraq (ID:MT554100.1) the identical 96 %, and the clusters including HCV subtype 4a isolate AASN35 Iraq (ID:MT554099.1) the identical 99%.

Using genomics to examine the persistence of Streptococcus pneumoniae serotype 19A in Ireland and the emergence of a sub-clade associated with vaccine failures

BackgroundStreptococcus pneumoniae serotype 19A remains a significant cause of invasive pneumococcal disease (IPD) in Ireland despite the successful introduction of a 13-valent pneumococcal conjugate vaccine (PCV13) in 2010 which reduced the overall incidence of IPD in children. MethodsInvasive Streptococcus pneumoniae serotype 19A isolates from the Irish reference laboratory between 2007–08 and 2017–18 were analysed using whole genome sequencing (WGS) to investigate the persistence of this vaccine-preventable serotype. We compared the entire national 19A collection to other international collections using a standardised nomenclature of Global Pneumococcal Sequencing Clusters (GPSC). ResultsExpansion of GPSCs and clonal complexes (CCs) may have been associated with vaccine introduction and antimicrobial prescribing policies. A sub-clade of GPSC1-CC320 (n = 25) unique to Ireland, included five of the ten vaccine failures/breakthrough cases identified (p = 0.0086). This sub-clade was not observed in a global GPSC1-CC320 collection. All isolates within the sub-clade (n = 25) contained a galE gene variant rarely observed in a global pneumococcal collection (n = 37/13454, p < 0.001) nor within GPSC1-CC320 (n = 19/227) (p < 0.001). The sub-clade was estimated to have emerged at the start of the PCV-vaccine era (ancestral origin 2000, range 1995–2004) and expanded in Ireland, with most isolated after PCV13 introduction (n = 24/25). ConclusionsThe identification of a sub-clade/variant of serotype 19A highlights the benefit of using WGS to analyse genotypes associated with persistence of a preventable serotype of S. pneumoniae. Particularly as this sub-clade identified was more likely to be associated with IPD in vaccinated children than other 19A genotypes. It is possible that changes to the galE gene, which is involved in capsule production but outside of the capsular polysaccharide biosynthesis locus, may affect bacterial persistence within the population. Discrete changes associated with vaccine-serotype persistence should be further investigated and may inform vaccine strategies.

Antimicrobial Effect of Toothpastes on Dental Caries

Dental caries is an infectious disease of teeth which end up in the destruction of dental hard tissue. Poor oral hygiene is one of the major factor for accumulation of microorganisms and their harmful effects. The current study was done on antimicrobial efficacy of different brands of toothpastes against organisms isolated from patients having dental caries and comparative analysis has been made between chemical and herbal based toothpastes. Primary screening of organisms had been carried out on sterile Nutrient Agar medium and five isolates (A1, A2, A3, A4 and A5) were obtained. Antimicrobial activity of toothpastes was checked using Agar well diffusion assay. In chemical based toothpastes, Close-up showed the maximum zone of inhibition of 30mm against A1 isolate and minimum efficacy was observed with Pepsodent with zone of 7mm against A4 isolate. In herbal based toothpastes, Babool showed highest effectiveness with zone of 28mm against A3 isolate and minimum with zone of 10 mm with Dabur red and Himalaya against A1 and A3 isolate. The comparative analysis showed that chemical based toothpastes are more effective than herbal based toothpastes. Further studies on probiotic approach should be done which has been a popular method used to affect oral microbial community.

Isolation and identification of crude oil-degrading yeast strains from Khafji oil field, saud Arabia.

Twenty five yeasts isolated were isolated from Khurais oil field in Saudi Arabia and assayed to evaluate their biodegradability. Only five isolates (namely, A1, A2, A3, A4 and A5) showed potential use of oil as sole carbon source. During incubation period, highest growth rate were recorded for A1, A2 and A3 isolates. Low growth distinguished A4 isolate; A5 isolate could not degrade oil.Spectrophotometrical analysis for four yeast isolates biodegradation activities indicated that, A1 isolate was superior for oil degradation (61%) comparing with A4 isolate which reflected lowest degradation % (33%). A2 and A3 isolates showed moderate biodegradation activity (56 and 51% respectively).D1/D2 domain of the 26S rRNA gene sequence was used as molecular marker to identify five yeast isolates. After comparing 26S rRNA gene sequences of five yeast isolates with highly similarity isolates, five yeast isolates (A1, A2, A3, A4 and A5)were submitted to database as Candida tropicalis (MW488263), Candida tropicalis (MW488264), Rhodotorula mucilaginosa (MW488265) and Rhodosporidium toruloides (MW488266) respectively. Using OXF1/ACR1 primer, specific lipase gene amplicon with 250 bp were detected with in all four yeast isolates.

Research Note: Disease tolerance/resistance and host immune response to experimental infection with Pasteurella multocida A:1 isolate in Indian native Nicobari chicken breed

Indigenous chicken breeds are considered to be more disease tolerant than exotic chicken breeds especially for the bacterial diseases. Nicobari and Vanaraja chicken were evaluated for the survivability/mortality patterns and host immune response after experimental infection with P. multocida A1 isolate. The birds were inoculated with 1.9 × 105 CFU/mL through intraperitoneal (I/P) and intranasal (I/N) routes at 2 different age groups viz., 12 wk and 18 wk. Symptoms, mortality rates, lesions in dead birds were observed; Serum from surviving birds of different groups from both breeds were collected at 5, 14, 21, 28, 35 and 42nd d and specific antibody titers were measured by indirect ELISA. At 12 wk of age, the mortality rates were 100% and 16% in birds inoculated by I/P and I/N routes respectively in Vanaraja birds; whereas the mortality rates were 50% and 16% I/P and I/N routes respectively in Nicobari birds. At 18 wk of age the mortality rates were 16% and 50% for I/P routes in Nicobari and Vanaraja birds respectively. The mortality rates were 16% for I/N route in both Nicobari and Vanaraja birds. Lesions such as necrotic foci on liver, congestion in the liver were observed in dead birds. Serum titers were significantly (P < 0.05) higher in surviving Nicobari birds inoculated through I/P route followed by I/N route. The peak titers were reached on 14th d postinfection and declined thereafter. However, no significant difference was found in I/N route of inoculation between 2 breeds. Nicobari chicken breed showed significantly higher survivability and longer mean death time than Vanaraja germplasm to experimental Pasteuralla infection at both the ages however the survivability rate in both breeds improved at later ages.

Capsular type diversity of Mannheimia haemolytica determined by multiplex real-time PCR and indirect hemagglutination in clinical isolates from cattle, sheep, and goats in Spain

This study compares the utility of a commercially available multiplex q-PCR assay for serotyping A1, A2, and A6 M. haemolytica serotypes with indirect hemagglutination, for determining the relative distribution of M. haemolytica capsular types associated with respiratory disorders in cattle, sheep, and goats. For the 129 isolates analyzed, both q-PCR and IHA assays exhibited nearly complete agreement for capsular types A1 (k = 0.965) and A2 (k = 0.888) and substantial agreement for A6 (k = 0.801). Despite the overall good performance of the commercial q-PCR, its effectiveness differed between the host origin of the isolates. The serotype was identified by q-PCR in 83.3 % of cattle, 77.8 % of goat, and 53.8 % of sheep isolates. Combining the results of both methods, A1 was the most prevalent in cattle and sheep (55.6 % and 22.25 %, respectively) but was not detected in goats, A2 was the most prevalent in goats (61.1 %) and the second most prevalent in cattle (16.7 %) and sheep (20.5 %). The prevalence of A6 was 7.4 %, 5.1 %, and 16.7 % in cattle, sheep, and goats, respectively. Other capsular types determined exclusively by IHA were A16 in cattle, A9 in goats, and A7, A8, A9, and A13 in sheep. Capsular type diversity was greater in sheep (H = 0.601) than in cattle (H = 0.408) and goat (H = 0.330) isolates. The commercial multiplex q-PCR is a valuable tool, alternative to IHA, for identifying isolates of capsular types A1, A2, and A6, the most frequent serotypes of M. haemolytica associated with respiratory disease in ruminants. However, when testing sheep isolates it should be complemented with immunological assays due to the wider range of serotypes implicated.

Phenotypic and Molecular Characterization of Penicillin and Macrolide-Resistant Streptococcus pneumoniae Serotypes Among Pediatric Patients in Addis Ababa, Ethiopia.

BackgroundIn several countries, introduction of the pneumococcal conjugate vaccine (PCV) has led to a decline in antimicrobial-resistant pneumococcal disease but has also resulted in a concomitant increase in antimicrobial-resistant, non-vaccine serotypes of Streptococcus pneumoniae. We sought to determine the magnitude of penicillin and macrolide resistance among pneumococcal serotypes and the mechanisms of macrolide resistance in Ethiopia, 5 years after the introduction of PCV10 in the country.MethodsSusceptibility to penicillin and erythromycin of 119 pneumococcal isolates collected from pediatric patients aged 0–15 years in Addis Ababa, Ethiopia, was tested using disc diffusion, and minimum inhibitory concentration (MIC) was also determined by Etest. Pneumococcal serotypes were determined by sequencing the cpsB gene and using Quellung reaction. Polymerase chain reaction (PCR) and PCR-restriction fragment length polymorphism analysis were used to detect and differentiate the macrolide resistance genes erm(B), mef(A), and mef(B).ResultsAmong the 119 isolates, 2.5% (3/119) were resistant to penicillin, while 58% (69/119) were intermediate. Resistance to erythromycin was observed in 33.6% (40/119) of the isolates with the highest level of resistance among isolates from middle ear discharge, i.e., 53.3% (8/15). Half (19/40) of the erythromycin resistant isolates were serotype 19A and among serotype 19A isolates, the majority i.e., 54.3% (19/35) were resistant to erythromycin. The most common macrolide resistance determinant was mef(E) with a prevalence of 50% (20/40).ConclusionFive years after introduction of PCV10 in Ethiopia, we observed that the prevalence of penicillin-resistant S. pneumoniae was low. However, there was a high level of macrolide resistance which was mostly in serotype 19A, and the resistance was mainly mediated by efflux pumps. Introduction of PCV13 (which covers serotype 19A) would significantly improve coverage of the macrolide-resistant serotypes. Continued surveillance of pneumococcal serotype distribution and their antibiotic resistance pattern in Ethiopia is warranted.

HPV73 in cervical cancer and distribution of HPV73 variants in cervical dysplasia.

HPV73 is classified as possibly oncogenic and is not recognized by most commercial primary HPV screening platforms. The aim was to determine the prevalence of HPV73 among invasive cervical cancers, formalin-fixed paraffin embedded (FFPE) samples (N = 69), from southern Sweden during 2009 to 2010. Another aim was to determine proportions of HPV73 among Aptima HPV assay negative cervical cancers (N = 9, out of 206 cancers) and of high-grade cytological cervical diagnosis (N = 75, out of 5807 high grade lesions) in liquid-based cytology (LBC) samples collected between 2016 and 2019. We also investigated the distribution of HPV73 variants A1, A2 and B among HPV73-positive cases. HPV73 was detected by multiplex MGP-PCR and Luminex, and HPV73 variants were identified by sequencing PCR amplicons. HPV73 was detected in 2.9% (2/69, 95% CI: 0.18-9.9) of the FFPE cervical cancer series. Among the Aptima HPV-negative LBC samples, HPV73 was present in 55.5% (5/9) of the cancers and 29.3% (22/75) of the different grades of cervical diagnosis. The A1, A2 and B variants were present in 6.9% (2/29), 82.7% (24/29) and 10.3% (3/29) of the HPV73-positive women, respectively. Among the seven HPV73 cancer cases (two FFPE samples and five LBC samples), six A2 and one A1 isolate were detected. In summary, the A2 variant of HPV73 was most common in our region. In addition, the observed prevalence of HPV73 (2.9%) in cervical cancers and its relative high occurrence (55.5%) among Aptima HPV-negative cancers urge that detection of HPV73 should be included in future primary HPV screening programs.

Evaluation of mild strain cross protection in cacao – further evidence of the protective potential of cacao swollen shoot virus strain N1 against the New Juabeng (1A) isolate under field conditions

The yield, canopy status and cocoa swollen shoot virus disease (CSSVD) symptom severity were assessed in cacao trees in a long-term field experiment originally established to study cacao swollen shoot virus strain 1A (CSSV-1A) epidemics in plots inoculated with the mild strain CSSV-N1. Yield of cacao was 73.7% and 32.1% lower in cacao plants inoculated with CSSV-1A and the non-inoculated respectively, when compared with cacao plants inoculated with CSSV-N1. CSSVD severity and canopy cover of cacao plants were significantly (P < 0.0001) affected by varying the number of inner rows inoculated with CSSV-N1 as well as individual CSSV strains. The treatments in which all the test cacao trees or first five inner rows received mild strain cross protection (T4 and T1) had the best canopy and lowest disease severity. Trees with CSSV-N1 protection had better canopy and lower CSSVD severity in comparison with the non-inoculated and CSSV-1A inoculated trees. DNA fingerprinting analysis indicated that genetic make-up of the planting material does not appear to contribute significantly to variation in disease severity or canopy status. Significant relationships were established between some soil parameters and CSSVD symptom severity (organic carbon, phosphorus, calcium, nitrogen, magnesium) as well as canopy status (phosphorus, potassium, magnesium).

Molecular characterization of Mannheimia haemolytica associated with ovine and caprine pneumonic lung lesions

This study investigated via polymerase chain reaction (PCR) three main serotypes (A1, A2, and A6) and nine virulence-associated genes in 71 ovine and caprine Mannheimia haemolytica isolates obtained from lungs (n = 349) with pneumonic lesions from a slaughterhouse in Iran. The lung specimens were collected from sheep (n = 197) and goats (n = 152) between December 2018 and January 2020. A total of 71 M. haemolytica isolates were identified in sheep (37/197; 18.8%) and goat (34/152; 22.4%) pneumonic lungs. Serotypes A2 (30/71; 42.3%) and A6 (29/71; 40.9%) were the most frequently detected, whereas the A1 serotype was detected with a frequency of less than 10% (7/71; 9.9%) and five isolates remained unknown. The virulence genes lkt, pomA, and nanH were present in all the isolates. The detection rates for the remaining virulence-associated genes were: gcp (95.8%), lpsA (93%), fhaC (90%), irp (70.4%), hf (57.7%), and sodC (21%). The sodC gene was exclusively detected among A2 isolates (50%), while the irp gene was more prevalent among A2 isolates and the hf gene among A1 and A6 isolates. These data may be useful for the typing of isolates in epidemiological studies. This study provides information about the main serotypes and the prevalence of virulence-associated genes among M. haemolytica ovine and caprine isolates in Iran.

ISOLATION OF PROTEIN FROM THE SPINE VENOM OF PTEROIS VOLITANS FOUND IN THE INDONESIAN OCEAN, USING A HEATING PROCESS, FOR ANTICANCER, ANTIRETROVIRAL, ANTIBACTERIAL, AND ANTIOXIDANT ASSAYS

Objective: This research investigates the antibacterial, anticancer, antioxidant, and antiretroviral activities of the lionfish spine poison extract.&#x0D; Methods: Isolation and purification of the phospholipase A2 (PLA2) protein obtained from the spine poison were conducted through the following stages, including, extraction of the venom by sonication, heating, and purification using gradual saturation levels of ammonium sulfate. Furthermore, the purity and concentration of PLA2 were analyzed using the Lowry test and Marinetti’s method, respectively, while its protein content was ascertained through SDS-PAGE. Toxicity was then evaluated employing the brine shrimp lethality test (BSLT), and its anticancer activity was assessed in human cervical carcinoma cells (HeLa cells). Finally, its antioxidant, antibacterial, and antiretroviral activities were analyzed using the DPPH method, agar diffusion test against Salmonella sp. and E. coli, and SRV-2 and RT-qPCR tests, respectively.&#x0D; Results: The protein demonstrated 37.79% inhibition for anticancer activity, IC50 1312 ppm for antioxidant activity, 98.81%, and 89.28% inhibition of E. coli and Salmonella sp. respectively for antibacterial activity and 98.13% inhibition for antiretroviral activity.&#x0D; Conclusion: It can be concluded that lionfish (Pterois volitans) has the potential to be developed as an antioxidant, anticancer, antibacterial, and antiretroviral agent. Furthermore, the pharmacological activity of its spine venom was determined by isolating PLA2 protein from its extract, using an optimum heating temperature of 70 °C and an ammonium sulfate saturation level of 80%.

Public health and clinical implications of multidrug-resistant shigellosis cases in a reception centre for refugees/migrants, Greece, October-December 2019.

From October to December 2019, 18 shigellosis cases (median age: 4.3 years, range: 0-21) were identified in the Reception Center for refugees/migrants on the Greek island of Samos. Fifteen cases (83.3%) were Afghani. Median time from arrival to symptoms onset was 53 days (40-101). Isolates from 14 cases, serotyped as Shigella flexneri 1b, and from three cases, serotyped as S. sonnei phase S (I), presented a multidrug-resistant phenotype. S. flexneri 1b isolates also produced extended-spectrum β-lactamases. Shigella flexneri 2a isolate from the remaining case was resistant to sulfomethoxazole, trimethoprim and pefloxacin. Improvement of hygiene and strengthening of laboratory investigation is needed.