Hypopygiopsis violacea, a species of fly of forensic importance, was recovered from a corpse and described for the first time. The morphological structures of the second and third instar larvae of four specimens were examined using light microscope. Observations were focused on three main morphological characters: cephalopharyngeal skeleton, anterior and posterior spiracles. Cephalopharyngeal skeleton of second instar larva is darkly pigmented and without accessory sclerite below the mouth hook. The anterior spiracles of second and third instar larvae have 8-9 papillae each, arranged in a single row. The posterior spiracle of second instar larva has two spiracular slits with no thickening of peritreme. This differentiates it from the third instar, whereby the latter has three slits for each posterior spiracle. Cephalopharyngeal skeleton of third instar larva is heavily pigmented. An accessory sclerite is found below the hook part of third instar larva but is absent in second instar. Peritreme of the posterior spiracle of third instar larva is thick almost complete encircling a button. The intersegmental spines of the cuticular surface are dome-shaped and unicuspid. Third instar larva of this species is large with size approximately 15 mm long. These findings provide important identification features of immature stages of Hy. violacea which could be useful in forensic entomology.
Peritonitis still remains a serious complication with high rate of morbidity and mortality in patients on CAPD. Rapid and accurate identification of pathogens causing peritonitis in a CAPD patient is essential for early and optimal treatment. The aim of this study was to use 16S rRNA and ITS gene sequencing to identify common bacterial and fungal pathogens directly from the peritoneal fluid without culturing. Ninety one peritoneal fluids obtained from 91 different patients on CAPD suspected for peritonitis were investigated for etiological agents by 16S rRNA and ITS gene sequencing. Data obtained by molecular method was compared with the results obtained by culture method. Among the 45 patients confirmed for peritonitis based on international society of peritoneal dialysis (ISPD) guidelines, the etiological agents were identified in 37(82.2%) samples by culture method, while molecular method identified the etiological agents in 40(88.9%) samples. Despite the high potential application of the 16S rRNA and ITS gene sequencing in comparison to culture method to detect the vast majority of etiological agents directly from peritoneal fluids; it could not be used as a standalone test as it lacks sensitivity to identify some bacterial species due to high genetic similarity in some cases and inadequate database in Gene Bank. However, it could be used as a supplementary test to the culture method especially in the diagnosis of culture negative peritonitis.
Different miRNAs are involved in the life cycles of Schistosoma japonicum. The aim of this study was to examine the expression profile of miRNAs in individual S. japonicum of different sex before and after pairing (18 and 24 dpi). The majority of differential expressed miRNAs were highly abundant at 14 dpi, except for sja-miR-125b and sja-miR-3505, in both male and female. Moreover, it was estimated that sja-miR-125b and sja-miR-3505 might be related to laying eggs. sja-miR-2a-5p and sja-miR-3484-5p were expressed at 14 dpi in males and were significantly clustered in DNA topoisomerase III, Rap guanine nucleotide exchange factor 1 and L-serine/L-threonine ammonia-lyase. Target genes of sja-miR-2d-5p, sja-miR-31- 5p and sja-miR-125a, which were expressed at 14 dpi in males but particularly females, were clustered in kelch-like protein 12, fructose-bisphosphate aldolase, class I, and heat shock protein 90 kDa beta. Predicted target genes of sja-miR-3483-3p (expressed at 28 dpi in females but not in males) were clustered in 26S proteasome regulatory subunit N1, ATPdependent RNA helicase DDX17. Predicted target genes of sja-miR-219-5p, which were differentially expressed at 28 dpi in females but particularly males, were clustered in DNA excision repair protein ERCC-6, protein phosphatase 1D, and ATPase family AAA domaincontaining protein 3A/B. Moreover, at 28 dpi, eight miRNAs were significantly up-regulated in females compared to males. The predicted target genes of these miRNAs were significantly clustered in heat shock protein 90 kDa beta, 26S proteasome regulatory subunit N1, and protein arginine N-methyltransferase 1. To sum up, differentially expressed miRNAs may have an essential role and provide necessary information on clarifying this trematode's growth, development, maturation, and infection ability to mammalian hosts in its complex life cycle, and may be helpful for developing new drug targets and vaccine candidates for schistosomiasis.
Malaria is a life-threatening disease caused by the Plasmodium sp. parasite. Infection results in heightened pro-inflammatory response which contributes to the pathophysiology of the disease. To mitigate the overwhelming cytokine response, host-directed therapy is a plausible approach. Glycogen synthase kinase-3β (GSK3β), a serine/threonine kinase plays a pivotal role in the regulation of inflammatory response during pathogenic infections. The present study was conducted to investigate the chemo-suppressive and cytokine-modulating effects of insulin administration in malaria-infected mice and the involvement of GSK3β. Intraperitoneal administrations of 0.3 and 0.5 U/kg body weight insulin each for four consecutive days into Plasmodium berghei NK65 (PbN)-infected mice resulted in chemo-suppression exceeding 60% and improved median survival time of infected mice (20.5 days and 19 days respectively compared to 15.5 days for non-treated control). Western analysis revealed that pGSK3β (Ser9) intensity in brain samples from insulin-treated (0.3 and 0.5 U/kg body weight) infected mice each were 0.6 and 2.2 times respectively than that in control. In liver samples, pGSK3β (Ser9) intensity from insulin-treated infected mice were significantly higher (4.8 and 16.1 fold for 0.3 and 0.5 U/kg bw respectively) than that in control. Insulin administration decreased both brain and liver pNF-κB p65 (Ser536) intensities (to 0.8 and 0.6 times for 0.3 U/kg bw insulin; and to 0.2 and 0.1 times for 0.5 U/kg bw insulin respectively compared to control). Insulin treatment (0.5 U/kg bw) also significantly decreased the serum levels of pro-inflammatory cytokines (TNF-α (3.3 times) and IFN-γ (4.9 times)) whilst significantly increasing the levels of anti-inflammatory cytokines (IL-4 (4.9 fold) and IL-10 (2.1 fold)) in PbN-infected mice. Results from this study demonstrated that the cytokinemodulating effects of insulin at least in part involve inhibition of GSK3β and consequent inhibition of the activation of NF-κB p65 suggesting insulin as a potential adjunctive therapeutic for malaria.
Methicillin-resistant Staphylococcus aureus (MRSA) is known to cause nosocomial infections and is now becoming an emerging problem in veterinary medicine. The objective of the study was to determine the presence of MRSA in 100 cats and dogs sampled between November 2007 and April 2008 at the University Veterinary Hospital, Universiti Putra Malaysia. MRSA was detected in 8% of pets sampled. Ten percent (5/50) and 6% (3/50) of the isolates were from dogs and cats, respectively. All MRSA isolates possessed the mecA gene and were found to be resistant to at least three antimicrobials with a minimum of Oxacillin MIC of 8 μg/mL. One isolate (CT04) had an extremely high MIC of >256 μg/mL. The MLST type ST59 found in this study have been reported earlier from Singapore and other countries as a strain from animal and community-associated MRSA respectively. Pulsed-field gel electrophoresis revealed five pulsotypes. Two isolates from cats (CT27 and CT33) and three isolates from dogs (DG16, DG20, and DG49) were respectively assigned to pulsotypes B and D. The study suggests that cats and dogs in Malaysia are potential reservoirs for MRSA.
Urinary Tract Infections (UTIs) consider as the most common infections worldwide, with higher risk in patients experienced Acute Appendicitis (AA). The purpose of this study was, to investigate the bacterial profile of UTIs in patients with non-ruptured AA postsurgically, and to assess age- and gender-related links of all AA cases in Karak region, Jordan. Urine samples obtained from 46 cases (32 male and 14 female) aged between 16-70 years were diagnosed as non-ruptured AA, following with isolation and characterization of isolated bacteria. Out of 46 AA cases, uropathogens isolated from 25 (54.3%) UTI cases. Out of these isolates; 42 (73.7%) were gram-negative isolates and 15 (26.3%) were gram-positive bacteria. The percentage of isolates were E. coli (26.3%), Enterobacter species (21%), Enterococcus faecalis and Klebsiella pneumoniea (10.5%) for each, Streptococcus saprophytics and Pseudomonas aeruginosa (7%) for each, Yersinia spp. and S. milleri (8.8%). Out of UTI cases, 20 cases (80%) possessed mixed culture, each of them had at least one of Enterobacterial species. i.e. Enterobacter spp. or E. coli or both. More precisely, out of all these positivecases, 2 cases had pure gram positive-bacterial infection (8%), while pure gram negative bacterial infection comprised 48% of them and the rest (44%) were mixed (gram-negative and gram-positive) bacterial infection. Moreover, study revealed a high prevalence rate of AA cases 24 (52.2%) in the ages of 16-22 years, then declining the rate with increasing the age, reaching the lowest rate (4.3%) in ages of 60-70. In addition to age factor, the males significantly more susceptible to AA cases than females by 2.2-fold. Antibiotic sensitivity test revealed high resistance capability of E. coli to the most used antibiotics except for nitrofurantoin. Bacterial isolates showing sensitivity against ciprofloxacin, trimethoprim/sulfamethoxazole, amoxicillin-Clavulanic acid and nitrofurantoin, with a superiority for the first two. Results demonstrate high prevalence rate of UTIs in patients with AA. For avoiding, the needless use of antibiotics through sticking to our accountability as healthcare provisioner to pursuit the antimicrobial management.
The organisms of the genus Leishmania are flagellated protozoan parasites and are the causative agents of leishmaniasis. This disease is a major health problem, especially in tropical countries. Currently, cutaneous leishmaniasis is treated by chemotherapy using pentavalent antimonials, but these drugs have serious organo-toxicity, drug resistance on several occasions, and low efficiency in controlling the infection. The present work is carried out to evaluate the in vitro antileishmanial activity of methanolic extracts and phytochemical fractions of two plants ethnobotanically used against leishmaniasis and skin infection, Calotropis procera and Rhazya stricta leaves against Leishmania major promastigote and amastigote stages and cytotoxicity against the Vero cell line. The leaves of C. procera and R. stricta were extracted with methanol and fractionated by petroleum ether, chloroform, ethyl acetate, n-butanol, and water. The methanolic extracts of the leaves of C. procera and R. stricta exhibited antileishmanial activity against L. major promastigotes with IC50 values of 66.8 and 42.4 µg mL-1, respectively. While their CC50 2.3 and 298 µg mL-1 and their SI 0.03 and 7.03 respectively. However, the fractionations of the methanolic extract of C. procera leaves revealed antiparasitic activity against both L. major promastigote and amastigote stages in vitro, which significantly increased with polarity with the exception of n-butanol. Hence the best activity was revealed by the water fraction (IC50 of 26.3 and 29.0 µg mL-1) for the two stages. In conclusion, further phytochemical investigation should be performed for the C. procera water extract in terms of antileishmanial active ingredient isolation that may enhance the possibility of avoiding toxic substances and overcome the low SI (1.1 and 1.01).
Much of the new research and investigation in pharmacy sciences are concerned with developing therapeutic agents, and identifying and finding new drugs with their chemical structure to treat different human diseases such as infectious diseases from natural products. Therefore, the present findings relate to isolating five new compounds the dichloromethane extract of Peperomia blanda (Jacq.) Kunth grown on Socotra Island, Yemen. two new secolignans; which have been proposed as peperomin I & J. These compounds were isolated together with the other two polyketides presented as surinone D and dindygulerione F. The chemical structures were elucidated and confirmed with nuclear magnetic resonance (NMR) and liquid chromatography-mass spectroscopy (LCMS) analysis. These compounds were first isolated and reported from this plant. These new compounds' antimicrobial activity has been evaluated, and minimum inhibitory concentration has been recorded in the range of 125-250 µg/mL. The pharmacotherapeutic spectrum of compounds was predicated using PASS software which showed potential activity.
Establishing a microbial diagnosis for patients with community-acquired pneumonia (CAP) is still challenging and is often achieved in only 30-50% of cases. Polymerase chain reaction (PCR) has been shown to be more sensitive than conventional microbiological methods and it could help to increase the microbial yield for CAP patients. This study was designed to develop, optimize and evaluate multiplex real-time PCR as a method for rapid differential detection of five bacterial causes of CAP namely Streptococcus pneumoniae, Burkholderia pseudomallei and atypical bacterial pathogens, Mycoplasma pneumoniae, Chlamydophila pneumoniae and Legionella pneumophila. Duplex and triplex real-time PCR assays were developed using five sets of primers and probes that were designed based on an appropriate specific gene for each of the above CAP pathogens. The performance of primers for each organism was tested using SYBR Green melt curve analysis following monoplex realtime PCR amplification. Monoplex real-time PCR assays were also used to optimize each primers-probe set before combining them in multiplex assays. Two multiplex real-time PCR assays were then optimized; duplex assay for the differential detection of S. pneumoniae and B. pseudomallei, and triplex assay for the atypical bacterial pathogens. Both duplex and triplex real-time PCR assays were tested for specificity by using DNA extracted from 26 related microorganisms and sensitivity by running serial dilutions of positive control DNAs. The developed multiplex real-time PCR assays shall be used later for directly identifying CAP causative agents in clinical samples.
Despite the medical, veterinary and forensic relevance of myiasis-causing flies, knowledge of their diversity in Saudi Arabia is limited especially in the southern region. Therefore, a survey of myiasis-causing flies in the Jazan region was carried out using Red Top Fly Catcher traps baited with either decomposing beef liver or a lure composed primarily from fishmeal during the period April 2013-March 2014. Twelve known species were identified and recorded in this study, seven species of them belonging to Calliphoridae (Chrysomya, Lucilia, and Hemipyrella) and five species belonging to Sarcophagidae (Sarcophaga). Two of these species were recorded for the first time for Saudi Arabia, namely Hemipyrella pulchra (Wiedemann, 1830) and Sarcophaga (Liosarcophaga) exuberans Pandellé, 1896. Images of the species recorded are also provided for the first time. The results expand the knowledge of geographical distribution, fauna, and habitat of the myiasis-causing flies in Saudi Arabia. Biological information and world-wide geographical distribution of these species are included together with some taxonomic remarks.
The present work aimed to identify camel ticks Hyalomma dromedarii and Hyalomma marginatum using direct sequence of the mitochondrial 16S rRNA gene and the detection of their natural infection rate with Rickettsia and Borrelia using the PCR/ hybridization method for amplification of the citrate synthase (gltA) gene. The phylogenetic analysis showed 99% similarity between Hyalomma dromedarii and its reference with accession # L34306.1, as well as between Hyalomma marginatum and its reference with accession # KT391060.1 obtained from GenBank data base. The prevalence of H. dromedarii and H. marginatum was about 99% and 1%, respectively. The intraspecific variation among H. dromedarii ranged between 0.2-6.6%. The interspecific variation between H. dromedarii and H. marginatum was 18.3%. PCR/hybridization of the sampled H. dromedarii detected about 31%, 37% and 18% natural infection with Rickettsia, Borrelia and co-infection with both pathogens, respectively. In contrast, none of Rickettsia or Borrelia was detected in H. marginatum. The present study emphasizes the accuracy of the identification of camel ticks based on molecular techniques. The ability of H. dromedarii to spread more than one disease is an important issue from the epidemiological standpoint. Future epidemiological research should be carried out in Saudi Arabia to monitor the distribution of tick species and suggest effective control strategies.
Leishmaniasis is a widely spread zoonotic disease caused by the bite of infected sandflies, particularly in developing countries. Cutaneous leishmaniasis can have a diverse range of presentations, ranging from minor skin nodules to significant mucosal damage. However, nose involvement is infrequent. Our report highlights a 15-year-old female patient with a persistent skin lesion on her nose for three months, which is a rare manifestation of cutaneous leishmaniasis. The lesion started as a raised spot with a brownish-red color and a crust but eventually developed into an ulcer that spread over the entire lobe of the nose and even moved toward the eye. Microscopic examination revealed the presence of Leishmania amastigotes, and a biopsy confirmed a diagnosis of cutaneous leishmaniasis. The patient received daily intravenous sodium stibogluconate doses of 9 mg/kg for 20 days, and three weeks later, there was a significant clinical improvement, with the ulcer beginning to heal and no more amastigotes visible on microscopic examination. It is crucial to keep cutaneous leishmaniasis in mind as a possible diagnosis for patients with skin lesions, even in regions where the condition is not prevalent.
The emergence of methicillin-resistant Staphylococcus aureus (MRSA) infection is one of the greatest threats to both animal and human health. Our investigation was aimed to identify and differentiate between MRSA and methicillin-sensitive Staphylococcus aureus (MSSA) recovered from mastitic milk using MALDI-TOF mass spectrometry compared with phenotypic methods and studying their susceptibility to various antibiotics. Four hundred milk samples from mastitic animals (cows, sheep, goats, and dromedary camels) were investigated. Phenotypic identification of S. aureus was made through MASTASAPH Latex test, STAPH ID 32, and Vitek 2 system. The proteomic characterization of S. aureus was done by MBT. The Kirby Bauer method was accomplished to detect the resistance of S. aureus strains to antibiotics. The results of the MASTASAPH Latex test, revealed that 54 (46%) were recognized as S. aureus. All S. aureus isolates were identified by MBT with a score of more or equal 2.00. Several peaks were identified in the mass of 4590 Da, 4863 Da, and 4938 Da for MSSA and in the mass of 2636 Da and 3009 Da for MRSA. The MSP dendrogram demonstrated that the S. aureus isolates were classified into one group with a distance level of less or equal 400. The percentage of S. aureus resistance against carbenicillin, erythromycin and kanamycin was 94.4%, 38.88%, and 33.33%, respectively. In conclusion, S. aureus bacteria are among the key triggers for mastitis in Saudi Arabia. MBT is reported to be not only the rapid tool to identify S. aureus but also able to discriminate MRSA from MSSA.
Anaplasma marginale is the most prevalent tick-borne haemoparasite of cattle and causes huge economic losses to the dairy industry worldwide. This study aimed to determine the occurrence of A. marginale infection in blood and tick samples collected from livestock animals in the districts located in Khyber Pakhtunkhwa (KPK), Pakistan. A total of 184 blood and 370 tick samples were included in this study. It has never been reported that sheep, goats, and cattle in Tank, Ghulam Khan, Birmil and Miran Shah areas were infected with A. marginale. All samples of blood and ticks were collected through random sampling from March 2021 to January 2022 from cattle, sheep and goats and screened through PCR for anaplasmosis by using primer pairs of Anaplasma spp. Three hundred and seventy ticks were collected from infested hosts (120/184, 64.21%). Among the four morphologically identified tick species, the highest occurrence was recorded for Rhipicephalus sanguineus (n=138, 37.29%), followed by Rhipicephalus microplus (n=131, 35.4%), Rhipicephalus annulatus (n=40, 10.81%), Hyalomma anatolicum (n=31, 8.37%), and Hyalomma marginatum (n=30, 8.1%). The occurrence of female tick was highest (n=160, 43.24%), followed by nymphs (n=140, 37.38%) and males ticks (n=70, 18.9%). Among these ticks, A. marginale was detected in female ticks of R. microplus, and R. sanguineus. Molecular identification of A. marginale was confirmed in 120 out of 184 blood samples and 6 out of 74 tick samples. Overall, occurrence of A. marginale in blood and tick samples was found to be 65.21% and 8.1% respectively. Species-wise occurrence in blood samples of goats were 71.11% followed by sheep 68.31% and cattle 50%. Specie-wise occurrence of A. marginale in tick samples of cattle were 12.5% followed by goats 6.89%. The obtained sequence showed similarity with A. marginale reported from Kenya and USA. We report the first PCR based detection of A. marginale infection in blood samples and in R. sanguineus ticks of goats simultaneously.
A total of 719 wild rats were captured from four localities representing the west (Kuala Lumpur), east (Kuantan), north (Georgetown) and south (Malacca) to determine the diversity of blood protozoan from the urban wild rat population in peninsular Malaysia. Five rat species were recovered with Rattus rattus diardii being the most dominant species, followed by Rattus norvegicus, Rattus exulans, Rattus annandalei and Rattus argentiventer. Two blood protozoan species were found infecting the rodent population namely, Plasmodium sp. (42.1%) and Trypanosoma lewisi (25.0%). This study reports the presence of Plasmodium sp. for the first time in the rodent population in Malaysia. Two main intrinsic factors were identified affecting the parasitic infections. Trypanosoma lewisi infections were influenced by host age and sex with infections observed higher in male and juvenile rats meanwhile Plasmodium sp. infections were observed almost similar in both sexes. However, infections were higher in sub-adult rats.
Knowledge of molecular identification of tick-borne pathogens in camels in Saudi Arabia is very limited; few molecular epidemiological studies have been under taken. This study was to detect Anaplasma spp. and Piroplasma spp. in camels from Asir Province, Saudi Arabia. A total of 150 blood samples were collected from camels in Asir Province and investigated by polymerase chain reaction (PCR) that targeted 18S rRNA and 23S rRNA to detect the DNA of Piroplasma spp. and Anaplasma spp., respectively. The positive samples for 23S rRNA were assayed again by PCR targeting the 16S rRNA. All the blood samples were free from Piroplasma spp. infection. Three camels (2%) were found to be positive for Anaplasma infection through use of PCR that targeted the 23S rRNA gene. There were no significant differences between ages or sexes in the camels that tested positive for Anaplasma. All positive Anaplasma infections were recorded in camels that were infested by ticks. Two Anaplasma sequences for the16S rRNA gene were deposited in GenBank with accession numbers MN882724 and MN882725. They recorded 99.16% and 99.34% similarities (respectively) with KF843825.1 (Candidatus Anaplasma camelii reported in Unizah, Saudi Arabia). Phylogenetic analyses revealed that the two sequences recorded in this study were close to each other; both were located in one cluster with Candidatus Anaplasma camelii isolates that were recorded before in the adjacent areas of Unizah in Saudi Arabia and Iran. In conclusion: two new Anaplasma genotypes close to Candidatus Anaplasma camelii were found in camels in Asir Province, Saudi Arabia for the first time. The camels in this province were found to be free of Piroplasma infection.
The early molecular identification of strains of Plasmodium vivax that have a worse prognosis is important to stratify the risk of complications and choice of conduct made by medical teams. Thus, the aim of the present study was to associate the presence of polymorphisms in the pvmdr-1 and pvcrt-o resistance genes of P. vivax in patients with better or worse prognosis. This cross-sectional epidemiological study was conducted based on data obtained from the records of 120 patients diagnosed with malaria in the Brazilian Amazon. The T958M and F1076L mutations of the pvmdr-1 gene had a frequency of 3.3 and 4.2%, respectively, and primo-infected patients had a 17 times greater chance of being infected with protozoa with the T958M mutation compared to patients with previous episodes. Regarding pvcrt-o, the C393T and T786C polymorphisms had a frequency of 14.2 and 3.3%, respectively, and self-declared white patients had a 3.1 times greater chance of being infected with protozoa with the C393T polymorphism. In addition, patients with this pvcrt-o polymorphism had lower concentrations of C-reactive protein, indicating a better prognosis. These data present clues of genetic indicators useful for assessing the virulence of the parasite and the prognosis of patients with vivax malaria.
A study was undertaken to evaluate the relevance of detecting IgM and IgG antibodies in diagnosis of canine leptospirosis in Kerala, a southern state of India, which is endemic for the disease. A total of 205 blood (35 from healthy vaccinated, 30 from healthy unvaccinated and 140 from diseased dogs) and 151 urine samples (11 from healthy vaccinated and 140 from diseased dogs) were collected from three districts of Kerala, Thrissur, Palakkad and Kozhikode with high incidence of leptospirosis. Recombinant LipL41 protein was used as antigen and IgG and IgM based ELISAs were standardized. The results were compared with the gold standard test, microscopic agglutination test (MAT). The MAT positive samples (146 samples) were divided into those having titre >1:800 and those between 1:100 and 1:400 in view that the former constituted the acute cases. It was found that IgM ELISA was more specific and sensitive in detecting acute cases (MAT >1:800) whereas IgG ELISA was less specific. In case of seroprevalence studies (MAT titre 1:100 to 1: 400), IgG ELISA was found to be more sensitive and specific than IgM ELISA. Receiver operating characteristic curves when plotted, revealed the accuracy of IgM ELISA in acute leptospirosis. Many samples were positive for both IgG and IgM antibodies. Polymerase Chain Reaction (PCR) targeting lipl41 gene was standardized and urine and blood samples from the same dogs were tested. PCR was found to be the specific test for the early detection of leptospires in blood even before seroconversion. However, PCR analysis of the urine samples was found to be insensitive. Hence, it can be concluded that the diagnostic strategies should be modified, and a combination of serological and molecular tests is recommended in endemic areas rather than simple detection of IgM or IgG antibodies, for the early detection of acute clinical cases of leptospirosis.
Hand, foot and mouth disease (HFMD) is a highly contagious viral disease that predominantly affects children younger than 5 years old. HFMD is primarily caused by enterovirus A71 (EVA71) and coxsackievirus A16 (CV-A16). However, coxsackievirus A10 (CV-A10) and coxsackievirus A6 (CV-A6) are being increasingly reported as the predominant causative of HFMD outbreaks worldwide since the past decade. To date, there are still no licensed multivalent vaccines or antiviral drugs targeting enteroviruses that cause HFMD, despite HFMD outbreaks are still being frequently reported, especially in Asia-Pacific countries. The high rate of transmission, morbidity and potential neurological complications of HFMD is indeed making the development of broad-spectrum antiviral drugs/agents against these enteroviruses a compelling need. In this study, we have investigated the in vitro antiviral effect of 4 Ganoderma neo-japonicum Imazeki (GNJI) crude extracts (S1-S4) against EV-A71, CV-A16, CV-A10 and CV-A6. GNJI is a medicinal mushroom that can be found growing saprophytically on decaying bamboo clumps in Malaysian forests. The antiviral effects of this medicinal mushroom were determined using cytopathic inhibition and virus titration assays. The S2 (1.25 mg/ml) hot aqueous extract demonstrated the highest broad-spectrum antiviral activity against all tested enteroviruses in human primary oral fibroblast cells. Replication of EV-A71, CV-A16 and CVA10 were effectively inhibited at 2 hours post-infection (hpi) to 72 hpi, except for CV-A6 which was only at 2 hpi. S2 also has virucidal activity against EV-A71. Polysaccharides isolated and purified from crude hot aqueous extract demonstrated similar antiviral activity as S2, suggesting that polysaccharides could be one of the active compounds responsible for the antiviral activity shown by S2. To our knowledge, this study demonstrates for the first time the ability of GNJI to inhibit enterovirus infection and replication. Thus, GNJI is potential to be further developed as an antiviral agent against enteroviruses that caused HFMD.
Is Acanthamoeba sp. normally found in the eyes? A study was carried out to establish the possibility of Acanthamoeba sp. as a part of the normal conjunctival flora. Conjunctiva swabbing were carried out in 286 healthy Orang Asli school children using sterile cotton swab. The swab was then inoculated onto non-nutrient agar (NN-A). Heat killed Escherichia coli that was used as food source for the growth of the amoebae was pipetted onto and away from the smear. The plates were incubated at 30 degrees C and examined daily using an inverted microscope for 14 days. Morphology of the trophozoites and cysts of the amoebae were used as the taxonomic criteria for identification. Positive-controls and negative-controls were done to check for the consistency of the technique used and monitoring of contamination respectively. None of the conjunctiva swab cultured was positive for Acanthamoeba sp. This finding may indicate that Acanthamoeba sp. is not part of normal conjunctival flora or conjunctiva swab is an insensitive technique to isolate the organism. However, a more extensive research is needed to investigate these possibilities.