Serological and molecular detection of neurocysticercosis among epileptic patients in Nagpur, Maharashtra state (India).

Neurocysticercosis (NCC), one of the most important neuroparasitic diseases in humans, is caused by Cysticercus cellulosae, the metacestode stage of digenetic zoonotic cestode Taenia solium. The present study aims at the detection of anti-cysticercus antibodies in the sera of epileptic patients (n=26) visiting a tertiary care hospital in Nagpur, Maharashtra state, India, by an in-house developed indirect IgG-ELISA and enzyme-linked immunoelectro transfer blot (EITB) assay using different antigens (namely, Whole Cyst Antigen (WCA), Cystic Fluid Antigen (CFA), Scolex Antigen (SA), Excretory-Secretory Antigen (ESA) and Membrane-Body Antigen (MBA)) prepared from T. solium metacestodes to find out the status of NCC. An attempt has also been made for molecular detection of NCC from blood samples of those patients by Polymerase Chain Reaction (PCR) assay targeted at large subunit rRNA gene of T. solium. The IgG ELISA level of anti-cysticercus antibodies against WCA, CFA, SA, ESA and MBA antigens were as follows: 19.23 %, 23.07 %, 38.46 %, 30.76 % and 15.38 %. The seroreactivity to CFA, SA and ESA was found in equal proportions in patients with ring-enhancing lesions. In the EITB assay, the lower and medium molecular weight protein bands of SA and ESA were immunodominant compared to the higher WCA and CFA peptides. PCR positivity could be observed in 34.6 % (9/26) of the patients under study. It is the first report of detecting NCC among epileptic patients of the Nagpur region of Maharashtra state in India using serological and molecular tools.

Association of Myroides odoratimimus in immunocompromized piglets with post weaning multisystemic wasting syndrome.

AIM: To study the association of opportunistic infection due to Myroides odoratimimus in piglets immunocompromised by porcine circovirus type 2 (PCV2) infection. METHODS AND RESULTS: The clinical samples (n = 101) were analysed bacteriologically. The isolates were identified by their phenotypes and MALDI TOF-MS analysis as Myroides species. The phylogram constructed based on nucleotide sequences of the 16S rRNA gene showed identity (~99%) with the M. odoratimimus isolates. The minimum inhibitory concentration values for antibiotics revealed M. odoratimimus to be resistant against carbapenem, cephalosporins, aminoglycosides and fluoroquinolones. The presence of PCV2 in affected tissue samples was confirmed by amplification of the 565 bp region of ORF2 of the PCV2 genome. The topology of the phylogenetic tree grouped the PCV2 with cluster-2d. CONCLUSIONS: PCV2 being immunosuppressive in nature might have impaired the immunity thereby increasing the susceptibility of immunocompromised piglets to opportunistic pathogens such as M. odoratimimus leading to disease severity and high mortality. The M. odoratimimus isolates were found to be multidrug resistant and evidenced for uncertain clinical relevance and hence could act as hidden source of public health hazard. SIGNIFICANCE AND IMPACT OF THE STUDY: Myroides odoratimimus is a rarely reported human pathogen. We reported the incidence of infection due to seemingly rare isolates of M. odoratimimus causing an outbreak of pneumonia in piglets. This appears, to the best of authors' knowledge, to be the first outbreak due to Myroides recorded in animal clinical cases described in the literature.

Distribution of Orientia tsutsugamushi in rodents and mites collected from Central India.

Orientia tsutsugamushi, the causative agent of scrub typhus, is an obligate intracytosolic bacterium transmitted among humans and small mammals by some species of larval trombiculid mites (chiggers). It has been recognized as a pathogen of major public health concern in the Asia-Pacific region. As disease is considered as a neglected, there exists a gap in our knowledge of the disease with regard to the sporadic epidemiologic data in endemic areas. The purpose of the study was to find out the vector as well as pathogen distribution in rodents present in the scrub typhus-reported areas in central India. We studied the seasonal variations of occurrence in O. tsutsugamushi in rodents and mites by molecular detection targeting the 56-kDa and 47-kDa genes. Rodent and mite samples were collected during December 2015 to July 2017. A total of 127 samples from rodents, seven pools of mites, and four pools of fleas were collected and processed for DNA isolation. Nested PCRs targeting the 56-kDa and 47-kDa surface antigen genes were performed. In addition, quantification of bacterial load was done by qPCR targeting the 47-kDa gene. During the pre-monsoon season, O. tsutsugamushi was detected in 12% and 10% samples employing the 56-kDa and 47-kDa nested PCRs, respectively, whereas, during post-monsoon season, the respective detection rates were 13.33% and 26.66%. This study predicted a bimodal pattern during the months of pre-monsoon and post-monsoon season with a peak in post-monsoon. Thus, the impact of season on the perpetuation of O. tsutsugamushi in the host was observed.

Brucellosis in migratory sheep flock from Maharashtra, India.

Brucellosis is a zoonotic disease worldwide distributed and having the economic as well as public health importance. The prevalence of brucellosis among sheep flock having history of abortions was studied. A total of 229 samples comprising of 157 blood and 72 clinical samples (vaginal swabs) were collected from 157 animals. Clinical samples were processed for the isolation of Brucella melitensis. Serum samples (n = 157) were tested by Rose Bengal plate test (RBPT) and i-ELISA. A total of 68 (43.31%) and 104 (66.24%) samples were positive by RBPT and ELISA, respectively. Brucella isolates (n = 2) were recovered from clinical samples. Both isolates demonstrated amplification for bcsp 31 and IS711 genes. On AMOS PCR, both the isolates amplified at 731 bp, i.e., belongs to B. melitensis species. The incidence of B. melitensis in a migratory flock warns the thorough testing and culling of Brucella-infected sheep from the flock on a continuous basis; otherwise, such incidence will be routine and poor farmers will be at a loss.

Isolation and characterization of multidrug-resistant Leclercia species from animal clinical case.

Leclercia adecarboxylata, a Gram-negative bacillus of family Enterobacteriaceae, is an uncommonly identified pathogen isolated from environmental and clinical specimens. Most of the human infections are polymicrobial and commonly occur in immunocompromised hosts, although nosocomial infections in immunocompetent hosts have been documented. Here, we describe the case of isolation of Leclercia species as polymicrobial infection from bovine suffering from respiratory distress in Chhattisgarh state of India. The isolates were identified by their phenotypes, 16S rDNA sequencing and MALDI-TOF-MS. The isolate was found to be resistant to aminoglycosides and fluoroquinolone antibiotics and intermediate resistant to cephalosporins and evidenced for uncertain clinical relevance and could act as hidden source of public health hazard. SIGNIFICANCE AND IMPACT OF THE STUDY: Leclercia adecarboxylata is a rarely reported human pathogen. We report here the case from bovine suffering from respiratory distress; the sample yielded Leclercia species as polymicrobial culture. The isolate was found to be multidrug resistant and evidenced for uncertain clinical relevance and could act as hidden source of public health hazard. The limited literature available on this organism is reviewed, and the potential implications of findings are discussed. To the best of our knowledge, this is the first report of isolation and characterization of multidrug-resistant Leclercia species from animal clinical case from India.

Isolation, antibiogram and pathogenicity of Salmonella spp. recovered from slaughtered food animals in Nagpur region of Central India.

AIM: To determine the prevalence, antibiogram and pathogenicity of Salmonella spp. in the common food animals slaughtered for consumption purpose at government approved slaughter houses located in and around Nagpur region during a period of 2010-2012. MATERIALS AND METHODS: A total of 400 samples comprising 50 each of blood and meat from each slaughtered male cattle, buffaloes, pigs and goats were collected. Isolation was done by pre-enrichment in buffered peptone water and enrichment in Rappaport-Vassiliadis broth with subsequent selective plating onto xylose lysine deoxycholate agar. Presumptive Salmonella colonies were biochemically confirmed and analyzed for pathogenicity by hemolysin production and Congo red dye binding assay (CRDA). An antibiotic sensitivity test was performed to assess the antibiotic resistance pattern of the isolates. RESULTS: A total of 10 isolates of Salmonella spp. from meat (3 from cattle, 1 from buffaloes and 6 from pigs) with an overall prevalence of 5% among food animals was recorded. No isolation was reported from any blood samples. Pathogenicity assays revealed 100% and 80% positivity for CRDA and hemolytic activity, respectively. Antimicrobial sensitivity test showed multi-drug resistance. The overall resistance of 50% was noted for trimethoprim followed by ampicillin (20%). A maximum sensitivity (80%) was reported to gentamycin followed by 40% each to ampicillin and trimethoprim, 30% to amikacin and 10% to kanamycin. CONCLUSION: The presence of multidrug resistant and potentially pathogenic Salmonella spp. in slaughtered food animals in Nagpur region can be a matter of concern for public health.

Humoral and delayed-type hypersensitive responses against listeria monocytogenes phosphatidylinositol-specific phospholipase C in experimentally infected buffaloes.

The kinetics of antibody production against phosphatidylinositol-specific phospholipase C (PI-PLC) and the isolation pattern of Listeria monocytogenes from bacteriological samples were studied following oral infection of buffalo calves with 3 x 10(9) cells each of pathogenic L. monocytogenes. Antibodies to PI-PLC appeared by 4-8 days post infection (PI), with a peak between days 7 and 16 PI, when tested by indirect plate-ELISA. Subsequently, antibody titres in all the animals declined and became undetectable on days 26-35 PI onwards until the study concluded on day 211 PI. Dot-ELISA could detect the antibodies to PI-PLC 1-2 days earlier and at higher titres as compared to plate-ELISA. L. monocytogenes could be recovered from faeces, nasal swabs and haemocultures from days 2 to 33, days 2 to 21 and days 11 to 17 PI, respectively. Antibodies to PI-PLC were detected during the course of active infection but their titres declined sharply once animals became culturally negative. Sonicated antigen elicited the highest delayed-type hypersensitivity response, followed by PI-PLC and listeriolysin O.

Isolation of pathogenic Listeria monocytogenes and detection of antibodies against phosphatidylinositol-specific phospholipase C in buffaloes.

The isolation of pathogenic Listeria spp. in bacteriological samples, and anti-phosphatidylinositol-specific phospholipase C (anti-PIPLC) antibodies in sera of buffaloes were studied. Isolation of the pathogen was attempted from the samples by selective enrichment in University of Vermont Medium and plating onto Dominguez-Rodriguez isolation agar. Pathogenicity of the isolates was tested by Christie, Atkins, Munch Petersen test and mice incoulation test. Listeria spp. and L. monocytogenes were isolated from 8.8 and 2.4%, and 4.8 and 1.6% of 125 each meat and blood samples, respectively. Out of the 125 samples each of feacal, nasal and vaginal swabs from buffaloes 8 and 4%, 13.6 and 2.4%, and 6.4 and 2.4% were positive for Listeria spp. and L. monocytogenes, respectively. L. ivanovii was confirmed from 0.8% vaginal sample. A total of 125 serum samples were tested by phosphatidylinositol-specific phospholipase C (PIPLC) based indirect ELISA of which 4.0% turned out to be seropositive.

Listeric infections in humans and animals in the Indian subcontinent: a review.

Listeriosis is an important bacterial zoonosis caused by the intracellular pathogen Listeria monocytogenes. The disease has been reported in animals from the Indian subcontinent, usually in the form of sporadic cases but occasionally as outbreaks. Cases of listeriosis arise mainly from the ingestion of contaminated food. Listeriosis has been reported to cause encephalitis, abortion, mastitis, repeat breeding and endometriosis in animals. Listeric infections occur in children and women with a poor obstetric history. The epidemiological aspects and pathogenesis of listeriosis in animals and humans are not yet fully understood. This review offers comprehensive information on experimental studies and field cases in animals and on cases of human listeriosis. There are also sections on isolation from foods, diagnosis and treatment in humans and animals.

The occurrence of pathogenic Listeria monocytogenes and antibodies against listeriolysin-O in buffaloes.

The occurrence of Listeria monocytogenes in meat and milk samples, and antilisteriolysin O (ALLO) antibodies in sera of buffaloes were studied. Isolation of the pathogen was attempted from the samples by selective enrichment in University of Vermont Medium and plating onto Dominguez-Rodriguez isolation agar. The pathogenicity of the isolates was tested by Christie, Atkins, Munch Petersen test and mouse inoculation test. Of 167 meat samples 2.4 and 10.17% were positive for L. monocytogenes and Listeria sp., respectively. Of the 64 milk samples 6.25 and 26.13% were positive for L. monocytogenes and Listeria sp., respectively. A total of 284 serum samples were tested by listeriolysin O (LLO)-based indirect enzyme-linked immunosorbent assay of which 25.35% were found to be seropositive. The culture positivity for L. monocytogenes and detection of ALLO did not show any agreement (kappa = 0.035). The prevalence of pathogenic L. monocytogenes in milk and meat and the occurrence of anti-LLO antibodies is of concern from the public health point of view.

Detection of anti-listeriolysin O and Listeria monocytogenes in experimentally infected buffaloes (Bubalus bubalis).

The kinetics of antibody production against listeriolysin O (ALLO) and the recovery pattern of Listeria monocytogenes from bacteriological samples were studied following oral infection of buffalo calves with 3 x 10(9) cells each of pathogenic L. monocytogenes. Antibodies to LLO appeared by 7-10 days post infection (PI), with a shallow peak between days 16 and 36 PI, when tested by indirect plate-ELISA. The titres of ALLO in all the animals then declined slowly but remained detectable up to day 70 PI. In dot-ELISA, ALLO could be detected by days 5 to 7 PI, and with higher titres than with the plate-ELISA. The pathogen was recovered at low rates as ALLO first appeared but was absent in the faecal, nasal and blood cultures as production of ALLO peaked.