Dataset: 11.1K articles from the COVID-19 Open Research Dataset (PMC Open Access subset)
All articles are made available under a Creative Commons or similar license. Specific licensing information for individual articles can be found in the PMC source and CORD-19 metadata.
More datasets: Wikipedia | CORD-19
Deep Learning Technology: Sebastian Arnold, Betty van Aken, Paul Grundmann, Felix A. Gers and Alexander Löser. Learning Contextualized Document Representations for Healthcare Answer Retrieval. The Web Conference 2020 (WWW'20)
Funded by The Federal Ministry for Economic Affairs and Energy; Grant: 01MD19013D, Smart-MD Project, Digital Technologies
The diagnostic methods based on PCR have the potential to be more sensitive and have a shorter turnover time, though they lack proper validation. These molecular tests have been designed for the detection of many virulence genes and are often the most sensitive methods for detecting them.
PCR-based methods for CPV infection in dogs have been shown to be more sensitive than traditional techniques. They have shown to detect very low concentrations CPV and feline parvovirus in unprocessed fecal samples, and have been more useful to diagnose and differentiate canine enteric pathogens as the definitive diagnosis is important primarily for epidemic control and prevention. The PCR assays avoid the necessity for culture for subsequent phenotypic tests and that when employed to a large number of diarrheic samples help in clarifying the role of a particular organism in the enteric disease.
In this study, all the diarrheic puppies (100%) and majority of the healthy non-diarrheic puppies (80%) were positive for C. difficile. A variety of 20 species of bacteria and 10 species of fungi were isolated from the rectal swabs taken from healthy dogs. Among them, E. coli, Streptococcus mitis, S. lactis, and Enterococci were more prevalent whereas Clostridium spp. and Lactobacillus spp. were least prevalent and neither Salmonella spp. nor Shigella were detected. Whereas, Clostridium spp. was most abundant in the fecal samples collected from dogs and cats (>20% on average). C. difficile was identified to cause of diarrhea in 10-21% of cases and postulated to be involved in some cases of acute hemorrhagic diarrhea syndrome in dogs. Thus, the increased prevalence of C. difficile in the fecal samples in puppies is explained by their normal inhabitance and abundance in the gastrointestinal tract of the puppies, more commonly among the diarrheic animals.
The prevalence of CPV-2b in the diarrheic puppies (90.3%) was more when compared with the healthy non-diarrheic puppies (10%). The shedding of CPV and CDV is strongly associated with acute hemorrhagic diarrhea. CPV-2 was typically seen in dogs without protective antibody titers, because of a lack of or an incomplete series of vaccinations. A window of susceptibility also occurs in puppies, in which maternal antibody falls below protective levels but vaccine-induced immunity is lacking. CDV was involved in acute hemorrhagic diarrhea without prominent respiratory and neurological signs.
Cpe, cpa toxin, STEC, E. coli – enterotoxin (LT), and CDV were identified in the puppies with hemorrhagic diarrhea, but none of them were isolated from the healthy puppies.
C. perfringens was responsible for a wide range of diseases in humans and animals. The pathogenicity of this species is associated with the toxin production such as the major toxins- alpha, beta, epsilon and iota, and the minor toxin-enterotoxin (cpe).
The identification of C. perfringens in the affected puppies could be related to the intestinal dysbiosis. The changes in the intestinal environment of dogs with diarrhea promote increased proliferation and transient overgrowth of enterotoxigenic strains of C. perfringens, leading to detectable amounts of enterotoxin (cpe) in the feces. Frequent growth of C. perfringens was observed in the intestine of dogs with CPV infection.
The main pathogenic elements of STEC and enterotoxigenic E. coli were classified into shiga toxin (stx1, stx2), heat-labile toxin (LT), and heat-stable toxin (ST). A higher percentage of diarrheic dogs were positive for hemolytic E. coli, with increased prevalence in young age as the intestinal epithelium appears to be more permeable than is the intestinal epithelium in older dogs.
Amoxicillin-clavulanate was recommended in patients with bacterial translocation. Gentamicin was identified as the drug of choice for treating Gram-negative gastroenteritis bacteria and parvoviral enteritis. This supports the maximum sensitive pattern of gentamicin in 95% of the puppies with HGE. Cefotaxime was found useful against few Gram-positive and most of the Gram-negative microbes implicated in parvoviral enteritis, which was in turn found to be sensitive in 20% of the affected puppies. The choice and response to antibiotics vary with each and individual animal, as the type and composition of gastrointestinal microflora are not similar in all the animals which explain the varying sensitivity pattern observed in the study.
There was no significant association between breed, gender, time to separation from dam, age of diagnosis and being a CD+ or CD- calf (P > 0.05). However, both breed and age of diagnosis had a P-value < 0.20 and were considered for the multivariable regression analysis. There were a significant higher proportion of calves having diarrhoea, and a higher proportion of calves that had died, in herds with CD+ calves than herds with CD- calves (P ≤ 0.01). CD+ calves came from larger herds (> 55 cows) than CD- calves (P < 0.001), and from herds where it was more common with calf group feeder (P < 0.05). The use of other antimicrobials than DHS to treat diarrhoea was more common in herds with CD+ calves (P < 0.001).
The median diversity of the E. coli isolates was 0.50 (50% CR: 0.23 - 0.72). There was no significant difference in diversity between samples from CD+ and CD- calves. Also, there was no difference in diversity between samples where the dominating PhP type was resistant to one or more antimicrobials and where the dominating type was not resistant.
Cow nutrition is closely associated with weak labor, amount of milk production, dystocia, and calf growth. Inadequate feed intake and macro- or micro-nutrient deficiencies during the last trimester increase calf morbidity and mortality rates because most fetal growth occurs during last 2 months of gestation. The quality and quantity of colostrum is associated with body condition score (BCS). A BCS near 5 (on a scale of 1~10) is acceptable for multiparous cows and a score of 6 for primiparous cows at calving is desirable. Recently, cow nutrition has been shown to impact the transition of the calf into adult life as well as fetal growth and development. Calves born to underfed cows have poor growth performance, low productivity, and higher susceptibility to disease. In another study, heifer calves born to cows fed supplemental protein during the last trimester were found to have greater pregnancy performance later in life compared to the control group.
Dystocia is closely related to poor calf performance as well as increased susceptibility to environmental pathogens which frequently cause calf diarrhea. Calves that experience dystocia may have physical symptoms such as congestion and swelling of the head and tongue, which can reduce the amount of colostrum uptake from the dam. The absorption rate of colostrum-derived immunoglobulin is lower in these calves compared to healthy animals. Consequently, the affected calves cannot obtain appropriate passive immunity from the dams due to inadequate colostrum uptake during early life (e.g., 2~ 6 h after birth).
The major causes of dystocia are associated with large calf size and small pelvic size of the dam. Large calves are more likely to have an improper position and presentation (e.g., backward, breech, and mal-positioned limbs or head) in the uterus. Under these conditions, the head and legs cannot enter the birth canal. Insufficient maternal pelvic size also can induce dystocia, especially in beef heifers. To prevent dystocia, the dam's genetic inheritance (e.g., adequate pelvic size and calving ease) should be taken into consideration during heifer selection, and frequent monitoring of the calving cow is required for appropriate calving assistance.
The average rainfall by month was divided into three groups, less than 275 mm, 275–375 mm and more than 375 mm in Table 4. The highest percentage (2458; 48.8%) of all the six diseases studied were related with less than 275 mm of average rainfall. 45% (201) malaria, 43% (539) enteric fever, 48% (413) diarrhea, 52% (312) encephalitis, 52% (708) pneumonia and 53% (285) meningitis fell under this category (chi-square p = 0.010). Average rainfall of 275–375 mm and > 375mm was associated with 24% (1203) and 27% (1373) of the total disease burden respectively. Analyzing the data of 2012, in six out of twelve months, there were ≤375 mm average rainfall (Fig 5). Higher incidences of encephalitis (p = 0.001) and meningitis (p = 0.001) happened while there was low rainfall. Incidences of diarrhea (p = 0.002), malaria (p = 0.001), pneumonia (p = 0.002) and enteric fever (p = 0.002) increased with rainfall, and then gradually decreased.
The avian gastrointestinal (GI) tract is home to complex and diverse bacterial populations that provide many beneficial functions to host, which includes conferring colonization resistance against the invading pathogenic microorganisms. Development of the GI microbiota in chickens occurs immediately after hatching and is influenced by both genetic and external factors like diet and environment (1). Unlike other animals, a newly hatched chick does not have acquired healthy maternal microbiota as they are housed separately from the adult hens immediately after hatch in commercial production (2). Therefore, the GI tract of newly hatched chickens is usually sterile and presents an empty ecological niche that provides easy access for the pathogen to colonize with limited restriction (2). This factor alone makes young chickens highly susceptible to enteric bacterial infections, such as Salmonella, which can result in different degrees of disease spectrum from a subclinical carrier state to a high mortality rate depending on the infecting bacterial serovar and host’s susceptibility.
Salmonella enterica subsp. enterica serovar Enteritidis is a zoonotic enteric pathogen that is most frequently associated with diarrheal disease in humans while chickens serve as asymptotic carrier (3). Consumption of contaminated eggs produced by infected layer hens is one of the leading causes of Salmonella food poisoning in humans (4). In chickens, S. Enteritidis can be easily transmitted horizontally via the fecal–oral route as well as vertically via the reproductive tract, which can contaminate the egg (5). Additionally, chickens can also harbor S. Enteritidis asymptomatically and persist throughout their lifespan, which makes the identification of infected chickens and the eradication of the pathogen much more challenging. Young chickens can be exposed to S. Enteritidis through numerous external sources like contaminated feed or environment. The sterile GI tract of the newly hatched chickens also provides ample opportunities for a pathogenic organism like S. Enteritidis to firmly establish its own niche in the gut as early colonizer and potentially further impact the development of the gut microbiota during the disease state. Early exposure to Salmonella in young chick could result in two potential alternative outcomes: high mortality rate or persistence of infection in surviving chickens (6). Prolonged persistent infection with S. Enteritidis in the GI tract of chickens throughout their lifespan could alter the development of gut microbiota and have detrimental effect on the overall gut health of the chicken host.
The impact of genetic background on the composition of chicken gut microbiota has been mostly investigated in broilers due to the association of intestinal microbiota with performance of broiler chickens in terms of feed conversion efficiency (7–11). Studies in broiler chickens have indeed shown evidence that host genotype had significant impact on shaping the composition of the gut microbiota (7, 9, 11). Few studies had explored the relationship between the host genotype and its influence on microbiota composition in layer chickens, especially related to disease resistance. The host genetic background plays an important role in the resistance and susceptibility to Salmonella infection (12). Several studies have reported that many genes have been found to be associated with Salmonella resistance in the chicken (6, 13). One of the key candidate genes, known as major histocompatibility complex (MHC), plays an important role in disease resistance in the chicken (13–20). University of California, Davis (UCD) maintains a number of congenic layer lines differing in MHC B-complex haplotypes. A study by Cotter et al. had previously examined the association of B-complex immunity to S. Enteritidis using 12 congenic lines from UCD, differing in various B-complex haplotypes (13). Results from the study had suggested that chickens from UCD254 (B15/B15) were more susceptible to Salmonella infection compared to other lines in term of mortality and morbidity (13). However, underlying mechanism associated with susceptibility to Salmonella remains to be elucidated. As microbiota is a significant contributor to disease resistance, two highly inbred line UCD254 (B15/B15) and UCD077 (B15/B16) at UCD were used to examine MHC effect on microbial community in chicken intestinal gut.
The main objective of this study was to examine the impact of host genetic background on influencing early establishment of microbiota in combination with S. Enteritidis infection to determine S. Enteritidis-associated alteration in gut microbiota.
Globally, 105 out of 109 (96.3%) herds were positive to at least one of the examined enteric pathogens (pathogenic E. coli, C. perfringens types A and C, TGEV, PEDV or RVA). Fifty-eight out of 109 (53.2%) submissions were positive for only one of these pathogens, 47 out of 109 (43.1%) were positive for more than one pathogen and, finally, 4 out of 109 (3.7%) were negative for all these agents.
E. coli strains were isolated from all submissions tested. However, only 11 of them were classified into defined pathotypes (ETEC and EPEC). Specifically, 9 isolates were classified as ETEC and 2 as EPEC (Table 2). Within ETEC isolates, 7 different virulence factor profiles were found; among ETEC strains, the most frequently detected virulence factors were STII (7 isolates), STI (7 isolates) and F4 (5 isolates). Both EPEC strains showed the same combination of virulence factors, Intimin and escV.
C. perfringens type A was isolated from 98 submissions (89.9%), which were all of them cpa positive by PCR. No isolation of C. perfringens type C was found, and all submissions resulted negative to cpb gene detection by PCR. The cpb2 toxin gene was found in 95 out of 98 (96.9%) C. perfringens type A isolates. By immunoblotting of these 95 strains, 20 isolates were β2 toxin strongly positive, 51 positive, 17 weakly positive and 7 negative. The result of production of α toxin by immunoblotting was positive in 34 isolates.
Regarding viruses, 47 out of 109 (43.1%) submissions were positive for RVA, 4 (3.7%) for PEDV and none of them for TGEV.
Table 3 summarizes all combinations of pathogens found in the studied diarrhoea cases. Noteworthy, pathogenic E. coli were only found in combination with other pathogens, and the maximum number of pathogens found in one submission was four (ETEC, C. perfringens type A, RVA and PEDV).
The emergence of the modern pig production, with more intensified farming, has been paralleled with an increase of neonatal piglet diarrhoea prevalence. Neonatal enteric problems in a herd are usually the result from the interaction of multiple factors that need to be examined to find rational means for intervention. Major determinants for the manifestation of neonatal diarrhoea include factors such as passive immunity transferred by colostrum and milk, environmental temperature and humidity, management and infection pressure by specific pathogens of the herd. The present study focused on different enteric pathogens able to cause neonatal diarrhoea in piglets in an important European pig producing country such as Spain, with specific focus on frequency of infections and co-infections. It was not possible, however, to address the specific role of other non-tested pathogens as well as non-infectious factors in the studied cases.
Both viral and/or bacterial pathogens were found in 96% of the submissions, which represents a fairly high number of cases with at least one infectious agent present. C. perfringens type A is considered rather an ubiquitous bacteria in the pig intestinal tract. Therefore, it is difficult to predict if it acted as primary pathogen or its multiplication was triggered by other infectious or non-infectious factors. In consequence, it was not possible to assess in which number of submissions the primary cause was a pathogen. Noteworthy, infectious diarrhoea in newborn piglets is usually related to the presence of a single pathogen and mixed infections are considered less common. However, almost half of the cases (43.1%) of the present study corresponded to mixed infections, with C. perfringens type A being present in all these cases.
Regarding viruses, RVA was the most frequently detected agent. Rotavirus is among the most prevalent pathogens in cases of neonatal diarrhoea, and is often detected as the sole infectious agent. In fact, several recent studies have described such high prevalence in pigs in Europe and North America [13; 14]. In contrast, only 4 submissions were positive to PEDV. After the 1980s, problems with PEDV in Europe declined and disease outbreaks have been occasionally seen during the last decades. In 2013, PEDV was introduced for the first time on the American continent and resulted in severe outbreaks of disease in the naïve population. Recently, PEDV isolates similar to the S-INDEL variants described in America have also been detected in Europe [35, 36]. Regarding the occurrence of PED in 2014–2015 recorded by the European Food Safety Authority (EFSA) Network, countries voluntarily reported 245 cases of pig herds meeting the PED case definition and 71 pig herds with RT-PCR confirmation of PEDV-genome; such PEDV-confirmed cases were found in Austria, Belgium, Spain, France, Italy, the Netherlands and Germany (EFSA, 2015). As expected, TGEV was not found in any of the submissions. TGEV is a cause of disease in most pig-producing areas of the world. However, outbreaks of TGE in Europe are rare, probably due to immunological cross-protection induced by PRCV, which is apparently ubiquitous in the continent.
ETEC has commonly been incriminated as the main aetiological agent of neonatal diarrhoea, although a recent survey conducted in Canada has suggested that the clinical importance of neonatal E. coli enteric problems have decreased during recent years. Pathogenic pathotypes of E. coli were found only in 10% of the submissions in the present study which may reflect such a decrease in prevalence. Anyway, it is known that the major pathotype of E. coli responsible for intestinal disease in pigs is ETEC and these data fit with the results of the current work, since 9 out of 11 E. coli pathotypes were ETEC (with major virulence factors being STII and STI toxins and F4 fimbriae). Curiously, two of the E. coli isolates consisted of EPEC strains, which are usually related with post-weaning diarrhoea. However, EPEC isolates from neonatal diarrhoea have been already described [38, 39]. In contrast, all tested submissions yielded non-pathogenic E. coli strains, which is in line with other published works. In a Danish study on neonatal diarrhoea in four commercial swine herds, non-haemolytic E. coli were the most predominant isolate obtained after aerobic culturing of both diarrhoeic and non-diarrhoeic piglets. This was also the case in a similar Swedish study on neonatal diarrhoea, where non-haemolytic E. coli was found in all piglets. The prevalence of classical porcine ETEC in both Scandinavian studies was very low in diarrhoeic piglets (less than 3%). This fact can be associated to modern swine production, in which pre-farrowing vaccination of sows against E. coli fimbriae antigens (F4, F5 and F6) is common.
C. perfringens type A was found in most of the submissions (89.9%), with detection of the cpb2 gene by PCR in the majority of cases. Moreover, in 88 cases such results were reinforced with the production in vitro of the β2 toxin by immunoblotting. This result fits well with a recent report from Poland, who found a 91.4% prevalence of C. perfringens type A at herd level. Such a high rate of detection raises the question of this bacterium as a true cause of diarrhoea or part of the microbiota, which may be up-regulated in enteric disease scenarios. In fact, the impact of α and β2 toxins on disease pathogenesis has not been conclusively answered. In contrast, C. perfringens type C was not detected in any of the samples. This bacterium causes disease in piglets in many areas of the world, but in a global perspective, it is considered much less important than other enteric pathogens. In the same Polish survey, for example, they found a 1.4% herd prevalence of this bacterium. Probably, during recent years, C. perfringens type C infections are rare in cases of neonatal diarrhoea due to sow vaccination. Indeed, in Spain, routine vaccination of sows pre-farrowing with beta toxoid vaccines is usual.
The distribution of enteric pathogens identified alone or in combination in the diarrheic and nondiarrheic foals up to 90 days of age is shown in Table 4. A total of 87% (49/56; p < 0.27) of the fecal samples from the diarrheic foals tested positive for enteric pathogens, in addition to 80% (48/60; p < 0.27) of those from the nondiarrheic animals. Among the diarrheic foals, 46% had coinfections, and 41% had monoinfections. In contrast, 47% of the nondiarrheic foals had monoinfections, and 33% had coinfections (p < 0.29) (Table 4). The most common coinfections involved Strongyloides westeri, Strongylus, and Salmonella spp.; combined infections with VapA-positive R. equi and C. perfringens toxin A were also common. However, no significant difference was observed between the most common enteric pathogens detected in the mono- versus coinfections (Table 5).
Eight foals died during the study (six within diarrhea group). Salmonella was detected in 7 out of 8 of the dead animals. The following coinfections were detected in the dead animals: (1) E. coli, Clostridium perfringens type A, Salmonella Panama, Strongyloides westeri, and Strongylus; (2) E. coli fimH, E. coli papC, Salmonella Infantis, and Strongyloides; (3) Salmonella Typhimurium and Strongyloides westeri; (4) Salmonella enterica subsp. enterica and Strongyloides westeri; and (5) Salmonella Muenchen and Strongyloides westeri (Table 6). Only one dead foal tested negative for all enteric pathogens. With regard to the two nondiarrheic foals that died, one was positive for E. coli and Salmonella Infantis, while the other was positive for E. coli fimH, E. coli ag43, Salmonella Newport, and Strongyloides westeri (Table 6). The relationships between the mortality rate and age in the diarrheic and nondiarrheic foals are shown in Table 6.
A one-site herd of 400 sows was the subject of Case 3. In July 2013, two boars were located close to the gestation unit. A week later, gestation sows showed anorexia (16.8%) and diarrhea (5.3%). Thereafter, in the gilts, diarrhea was evident in the nursery (3–7%) and fattener (5–23%). Two-day-old piglets showed watery diarrhea (100%) with a mortality rate of 95%. Affected piglets died from severe dehydration within two days of the onset of clinical signs. The course of the disease lasted approximately two months with an overall pre-weaning mortality of 50% during that period.
The results of applied statistical analysis revealed the existence of significant differences between the age of birds depending on their health status (healthy, PEC and PEMS). The Kruskal-Wallis test showed that the average age of healthy, PEC and PEMS turkeys differs significantly (P = 0.036). Multiple comparison test showed that this difference was only between healthy and PEMS turkeys (P = 0.03); healthy turkeys were about 7 weeks old and PEMS about 4 weeks old. Generally the older the turkeys were, the healthier they were. Such correlation was also implied by the independence chi-square test (P = 0.007, φ = 0.31). The calculated ORs of =3.57 and 3.75 indicated that the chance of PEC and PEMS symptoms in turkeys aged 1-4 weeks are above 3.5 times higher than the chance of such disease symptoms in the older group of 5-12-week-old animals. The OR =6.92 indicates that the possibility of PEMS relative to PEC symptoms in turkeys in the fattening phase is almost sevenfold higher than in turkeys over 13 weeks of age.
Based on the PCR assay, among the healthy puppies screened, 80% were found positive for C. difficile, and 10% were positive for CPV-2b.
Among the affected puppies, all were positive for C. difficile, 90.3% were positive for CPV-2b, 17.7% were positive for cpe, 9.7% were positive for cpa toxin, 6.4% were positive for E. coli shiga toxin (STEC), 6.4% were positive for E. coli – enterotoxin (LT), and 3.2% were positive for CDV (Figure-1). None of the puppies were shedding both the CPV and CDV together whereas, cpe together with cpa and LT were detected in two of the puppies with hemorrhagic diarrhea.
Salmonella sp., Campylobacter sp., and C. difficile toxin B, enteric CCoV and Rotavirus were found negative in 62 puppies with hemorrhagic diarrhea screened samples.
The fecal antibiotic sensitivity results revealed gentamicin to be sensitive in 95% of the cases, azithromycin in 50%, enrofloxacin in 25%, cefotaxime in 20%, and tetracycline in 5% of the cases. The order of sensitivity was gentamicin > azithromycin > enrofloxacin > cefotaxime > tetracycline. Maximum resistance (100%) to amoxicillin and least resistance (5%) to gentamicin were observed.
Calf diarrhea has been a major disease that negatively affects the cattle industry. The economic impact caused by this condition is significant although many new intervention strategies (e.g., vaccine, medications, and herd management) have been developed and implemented to minimize the economic loss. Persistence of this significant problem in the field may be attributed to the multifactorial nature of calf diarrhea including permutations of infectious diseases, a failure to clearly understand the disease ecology, poor environmental hygiene, and biased epidemiological data. Genetic diversity, continuous evolution, emerging pathogens, and/or environmental ubiquity of pathogens are factors that hinder effective control of the disease. Therefore, the genetic evolution of RNA viral pathogens such as BRV, BCoV, BVDV, BToV, BNoV, and Nebovirus should be kept in mind and monitored with regular genomic sequence updates. Non-group A BRV might be considered for future studies to increase the detection range of calf enteric pathogens. Emerging viruses should be regularly monitored for the evaluation of vaccines against calf enteric pathogens. Clinical significance of caliciviruses (BNoV and Nebovirus) must be carefully assessed to better control calf diarrhea in the future.
The use of highly sensitive diagnostic tests has increased the detection frequency of pathogens that were previously neglected. Therefore, optimized and appropriate diagnostic methods or platforms should be employed for detecting target pathogens in an accurate and timely manner with a minimum testing outcome bias. Currently, real-time PCR-based techniques are widely implemented in many veterinary diagnostic laboratories. These methods are highly accurate and provide high throughput performance but sometimes might overestimate the significance of pathogens detected in cases of calf diarrhea. The pros and cons of diagnostic test results and their overall interpretation must therefore be cautiously evaluated by referring clinical history from practitioner when the causative etiology is being determined.
Non-infectious risk factors have frequently been neglected by cattle producers, and also be considered equally important as infectious factors because the newborn animals are vulnerable to environmental stresses. The management and control of calf diarrhea before an outbreak is more cost-efficient than treating sick animals after the outbreak occurs. Although many enteric pathogens are involved in calf diarrhea, infection and transmission is accomplished via a fecal-oral route. Care must be thus taken to prevent pathogen transmission. Advice from professional consultants such as veterinarians and nutritionists is necessary to obtain an accurate diagnosis and control or manage risk factors associated with calf diarrhea in modernized large production systems.
In summary, the effective control of calf diarrhea should be based on three major points. First, a clear understanding of pathogen characteristics (e.g., mechanism underlying pathogenicity, prevalence in the field, and genetic evolution) is required. Second, advantages and disadvantages of various diagnostic methods and their application to diagnostic investigation along with clinical history should be considered. Finally, proper cow-calf management is necessary for disease prevention and control.
Case 2 involved a one-site herd of 350 sows with its own replacement gilts and the following parity distribution: 20% gilts, 40% parity 1–2, and 27.3% distributed amongst parity 3–5. Boars were purchased from a breeding company and were incorporated into the reproductive herd without quarantine. The farm was located within a few miles of a swine slaughterhouse in Buenos Aires. In February 2012, the pregnant sows showed anorexia (14–30%) and diarrhea (1%) associated with heat returns and abortions (3.3%). In the farrowing houses, approximately 100% of the lactating sows presented with anorexia. Pre-weaning mortality associated with the presence of diarrhea varied from 16.5% at the beginning of the outbreak to 27.9% 3 to 4 weeks after the initial clinical signs. An anatomopathological evaluation showed that 93.6% of the total pre-weaning mortality was due to diarrhea.
Sampling was conducted under the permission of the owners or other responsible persons. The birds in the farms were under the supervision of veterinarians, who took different samples as part of their routine work (i.e. as screening flocks for efficacy evaluation of applied vaccines or presence of any infections) and thus part of them were used in this study. For this reason, sampling did not require the approval of the Ethics Committee.
Considerable variations in the frequencies of the major enteric pathogens recovered from diarrheic and nondiarrheic foals worldwide have been reported in different studies [1, 6, 7, 13]. Moreover, many studies have focused on only one enteric pathogen [8, 9, 9, 10, 12, 40, 63, 64], despite the recognized etiological complexity and the potential combinations of pathogens in equine enteric infections [13, 52]. Notably, a large number of enteric pathogens of bacterial, viral, and parasitic origins, as well as diverse coinfections, have been identified in diarrheic and nondiarrheic foals. In fact, substantial etiological complexity of enteric organisms has been observed among foals, reinforcing the necessity to include major enteropathogens in the differential diagnosis of neonatal enteric infections in these animals [1, 7, 17]. The failure to identify significant differences in the frequencies of pathogens between diarrheic and nondiarrheic foals in the current study, with the exception of Clostridium perfringens toxin A+, has also been reported previously [6, 66], indicating that other factors, such as age, the nutrition and immune statuses, environmental conditions, general management practices, and characteristics of the farm facilities, may influence the occurrence of clinical cases of diarrhea.
In addition, no significant associations of specific coinfections with the presence (or not) of diarrhea were detected, and a high mortality rate was observed among the Salmonella-positive diarrheic foals, regardless of the serotype or other pathogens present. High mortality of foals with enteric salmonellosis has also been described elsewhere [1, 66]. These results provide important information for practitioners who detect Salmonella spp. in the feces of foals because proper antimicrobial therapy, rigorous hydration, and other critical care measures are required to prevent serious complications and to improve prognosis.
In our study, coinfection of at least two infectious pathogens is recorded in 2.4%, whereas higher prevalence was recorded by other researchers. In a study by Mesonero-Escuredo et al. recorded that 43.1% of samples were positive for more than one pathogen either bacterial or viral pathogens in pig neonatal diarrhea cases in Spain with C. perfringens type A being involve in all cases. However, Katsuda et al. noted that infectious diarrhea in newborn piglets is usually related to the presence of a single pathogen and mixed infections are considered less common. In another study by Zhang et al. recorded 4.3% of positive cooccurrences of enteric pathogens in sick children in Southwest China where E. coli and Norovirus were the predominant coinfective pathogens. These reports suggested that coinfection of pathogens varies from one region to another geographical region. Neonatal diarrhea, particularly in young piglets, is often complex with a mixture of infectious agents and other factors such as passive immunity transferred by colostrum and milk, environmental temperature and humidity, and managemental factor, all contributing to its manifestation.
In the present study, Picobirnavirus GG1 was recorded as highest cases as coinfecting agent. Picobirnaviruses are generally regarded as diarrheagenic viruses because most of their detection is associated with virus shed in feces and have been detected in animals and human patients with and without gastroenteritis and mostly coinfected with other enteric viruses such as Rotavirus, Astrovirus, Calicivirus, and Coronavirus.
Many researchers reported that RVA is the most common cause of viral diarrhea in neonatal pigs compared to Type C, and other workers have detected porcine Rotavirus in diarrheic fecal samples in nursing, weaning, and post-weaning pigs either alone or in combination of other enteric pathogens such as E. coli, Salmonella, and Adenovirus. Rotavirus A is only often detected as the sole infectious agent than as coinfection in cases of neonatal diarrhea. This is in agreement with our finding in which RVA was detected only in one diarrheic sample as coinfection with EPEC. Although it is clear that RVA has an overwhelming impact on diarrhea illnesses in children, coinfection with other enteric pathogens appears to aggravate diarrhea severity. These findings should serve as evidence for public health services when planning and developing intervention programs.
Diarrheagenic E. coli are important common bacterial pathogen involve in piglets diarrhea. According to Fairbrother and Gyles, the main pathotype of E. coli responsible for intestinal disease in pigs is enterotoxigenic E. coli and EPEC also known as attaching and effacing E. coli. Coinfection of EPEC with other infectious agents indicates that EPEC potentially accelerates gastroenteritis and is one of the main pathogens involved in coinfection with other diarrheagenic bacterial or viral pathogens. Shiga-toxigenic E. coli was detected as coinfective strain in two diarrheic samples. The strain is one of the leading causes of diarrhea in the developing world and importantly in Northeast India, and they are commonly recovered from diarrheic feces of food-producing animals including porcine.
Salmonellosis is one of the economically important enteric and septicemic diseases associated with morbidity and even mortality in farm animals. In our study, S. Typhimurium was the most frequent serovar involved in coinfection; however, this serovar is not host-specific serovar in pigs; hence, the finding is an indication of a public health and zoonotic concern. The close monitoring of such concurrent infection in porcine is important not only in terms of production but also from a public health point of view.
In the present study, higher detection of coinfection was recorded from unorganized farms, compared to organized farms. The reason for variation probably might be due to different management being practice from farm to farm, particularly in case of Picobirnaviruses which can enter and transmit mainly through sewage and untreated water in the farm. Crossbreed piglets recorded higher detection of mixed infection than local indigenous piglets. Although the variation is not very significant, it may be possible that the local non-descriptive piglets possess better immunity against natural infection than crossbreed animals. In addition, the weaning age of piglets of the local animals is generally 8-10 weeks in comparison with the crossbreed animals in organized farms (within 6 weeks). Maternal immunity might also play an important role in resisting the infection in piglets, particularly in indigenous pigs. In the same region in a study of diarrheagenic pathogens, reported a higher prevalence rate of Picobirnavirus and STEC in crossbreed than indigenous piglets population. In our study, samples collected during different seasons showed that coinfection of enteric pathogens was found to be most common during the summer (June-August). However, due to paucity of reports regarding seasonal variation of coinfection, hence, this did not allow us to compare our results. The humid climatic condition during summer with persistent rainfall in Northeast region may be an important reason for entry and transmission of infection among the pig population. Such climatic condition also allows suitable environment for persistent enteric pathogens such as E. coli, Salmonella, Rotavirus, and Picobirnavirus to cause diarrhea in piglets.
Diarrhea or scour has been recognized as one of the most common ailments causing deaths of young animals including piglets, but it is very often a neglected problem in most farming system. In the present study, a total of 11 diarrheic fecal samples were detected for a combination of at least two different enteric pathogens which are suggestive of the fact that a number of enteric pathogens may cause diarrhea either singly or in combination. Considering the importance of E. coli, Salmonella, Rotavirus, and Picobirnavirus as important diarrheal causing agents in animal species and human, hence, there is a possibility of transmission between porcine and human population, particularly in rural tribal areas in North East Hilly region of India were people reared two or three pigs in close proximity with human habitation and most share common drinking water.
Table 3 shows the association of the six diseases with the average humidity of the studied period of 2008 to 2012. We analysed the incidence of each disease in each of three groups of humidity recordings by month: less than 76.5%, 76.5–77.5% and more than 77.5% based on the mean humidity of the study site. Less than 76.5 percent humidity was associated with the highest percentage (2458; 49%; p = <0.01) of all the six studied diseases (Table 3). Average humidity of 76.5–77.5% and > 77.5% was associated with 27% (1373) and 24% (1203) disease respectively. However, in 2012, the average humidity was ≥ 76.5% in nine out of twelve months (Fig 4). Higher humidity was correlated with a higher number of cases of malaria (p = 0.0001), enteric fever (p = 0.0001) and diarrhea (p = 0.0001), but inversely correlated with meningitis (p = 0.0001), encephalitis (p = 0.0001) and pneumonia (p = 0.0001).
Environmental and maternal bacteria quickly colonize offspring gut after birth and shape theonset of a healthy intestinal immune system and its future development. Intestinal microbiota is characterised by its high population density, extensive diversity, and complexity of interactions throughout the gastrointestinal tract. Studies on the characterisation of the intestinal microbiota show that the major bacterial groups isolated from the pig intestine are Streptococcus, Lactobacillus, Prevotella, Selenomona, Mitsuokella, Megasphera, Clostridia, Eubacteria, Bacteroides, Fusobacteria, Acidodaminococci, and Enterobacteria. Interestingly, it was reported that there is clear evidence that gut microbiota play an important role in driving host metabolism and that the diversity of the faecal bacterial community and their changes over time were different in pigs depending on their subsequent susceptibility to post-weaning diarrhoea. The stomach and proximal small intestine (duodenum) contain relatively low numbers of bacteria (103–105 bacteria/g or ml of contents) due to low pH and/or rapid digesta flow. In contrast, the distal small intestine harbours a more diverse and numerically greater (108 bacteria/g or ml of contents) bacterial population.
The mean number of E.coli biochemical phenotypes in piglets increased as animals aged and E. coli populations in the pig faecal microbiota and in the farm environment are dynamic and show high levels of diversity.
Clinical manifestations of enteric colibacillosis obviously require the presence of pathogenic E.coli but also environmental changes and recognized risk factors. Moredo et al. demonstrated that the percentage of ETEC positive non-diarrhoeic pigs was 16.6% during the lactation period, 66% in the nursery phase and 17.3% in the finisher population. These data demonstrated that these pathogens can also be shed in faeces from healthy animals as already reported by Osek, in 1999.
The barrier functions of the gastrointestinal tract in the neonatal piglet is not as developed as in mature animals due to the higher pH in the stomach, the lower proteolytic capacity, and the dependence on passive immune protection due to immunological immaturity. The regeneration time of the small intestinal epithelium in day-old piglets is reported to be 7–10 days, as compared to 2–4 days in 3-week-old pigs. This difference is probably contributing to the susceptibility of infectious enteritis in new-born piglets, as a rapid turnover of enterocytes is considered a defence mechanism by the expulsion of infected cells. Taken together, all of these conditions combined with predisposing factors, contribute to the neonatal piglet’s vulnerability to ETEC enteric infections.
After weaning, the change in the intestinal environment of piglets, mainly due to dietary changes, results in an alteration of the composition of the indigenous flora. The diversity of E.coli strains of intestinal flora is usually high in healthy pigs, while in enteric colibacillosis we observe an alteration of the balance between the bacteria present in the normal intestinal flora. This condition leads to the proliferation of a dominating pathogenic strain, which colonizes the small intestine, rapidly reaching massive numbers to the order of 109/g of contents. This is the reason why frequently, if not always, samples collected in diarrhoeic pigs affected by colibacillosis allow the isolation of a pure culture of pathogenic E.coli.
This information must be considered for a correct interpretation of diagnostic results. In particular, the evaluation of diagnostic findings should be made only in consideration of both clinical signs and pathological lesions, while also taking into account the number of isolated pathogenic E.coli strains belonging to the identified pathotype and virotype.
Enteric disease outbreaks in pigs are frequently multifactorial. Focusing on the diagnosis and subsequent control strategies without take into account all the possible differential diagnoses or multiple agents involved can misguide practitioners. The diagnostic evaluation of neonatal and post-weaning colibacillosis requires similar standard diagnostic procedures. The diagnostic pathway is kept easy if the lesions observed during necropsy are strongly suggestive. This is the case in enteric colibacillosis, where bacteriology can usually easily confirm the suspect given by the recorded pathological lesions. Diarrhoea in the pre-weaned piglet is probably more straightforward to identify, treat, and prevent than post-weaning diarrhoea. In particular, ETEC neonatal diarrhoea must be differentiated from other causes of diarrhoea, such as Clostridium difficile, Clostridium perfrigens type A and C, enteric coronavirus (TGEV, PEDV) and rotavirus groups A, B and C. In piglets older than 7 days, coccidiosis due to Isospora suis should also be considered (Table 4).
ETEC PWD should be differentiated from other causes of diarrhoea already described in piglets such as EPEC, enteric coronavirus (TGEV, PEDV), rotavirus groups A, B and C, salmonellosis, proliferative enteropathy due to Lawsonia intracellularis and Brachyspira spp. (Table 5).
Diarrheic stool from 100 patients was collected. Fourteen samples were excluded from the molecular testing arm due to insufficient sample volume or sample processing deficiency. Patient demographics are outlined in Table 1. The mean age was 34.9 with 5 patients under the age of 14. 43.0% of patients were male, 80.2% lived in an urban area, 17.4% had access to well water only, 88.4% had access to proper latrines, and 81.4% practiced proper hand hygiene. Patients median duration of diarrhea was 10 days with 53.5% having acute diarrhea and 7.0% having chronic diarrhea. Median bowel movement frequency per day was 5. The median CD4 cell count was 361.5/μL with 36.1% having CD4 cell counts < 200/μL. The majority (72.1%) of patients were receiving highly active antiretroviral therapy (HAART) at the time of collection.
Out of the 100 specimens, 28 were positive for intestinal parasites using wet mounts and modified acid fast staining. C. parvum was the most common parasite at 28.6% followed by E. histolytica (17.9%), G. lamblia at (14.3%), C. cayetanensis (14.3%), S. stercoralis, (7.1%) and hookworm (3.6%).
The enteropathogens detected by the Allplex™ panel are outlined in Fig. 1. No failures were detected among the negative and positive controls. Only one patient had no organisms detected (when excluding B. hominis, D. fragilis and Aeromonas spp). The most common organisms detected were Shigella spp./enteroinvasive E. coli (EIEC) at 80.2%, enterotoxigenic E. coli (ETEC) at 73.3%, Aeromonas spp. at 73.3%, and enteroaggregative E. coli (EAEC) at 59.3%. Parasites detected included Blastocystis hominis (61.6%), Giardia lamblia (17.4%), Cryptosporidium spp. (10.5%), and Dientamoeba fragilis (8.1%). Viruses detected included norovirus GI/GII (16.3%), rotavirus A (4.7%) and adenovirus 40/41 (3.5%). Multiple pathogens were detected in 94.1% of stool specimens, with 64.0% having 5 or more enteropathogens (33.7% if excluding B. hominis, D. fragilis and Aeromonas spp.).
Older age was associated with Campylobacter spp., only occurring in patients between over the age of 35 (p = 0.04). D. fragilis was associated with increased frequency of diarrhea (median 4 movements when D. fragilis present compared to median 5, p = 0.02). Use of well water was associated with Cryptosporidium (33.3% with well water vs 6.0%, p = 0.001).
A box plot describing enteropathogen frequencies with CD4 cell count is outlined in Fig. 2. Only viral enteropathogens and STEC were associated with CD4 cell counts. A CD4 cell count < 500/μL was associated with the presence of a viral enteropathogen (p = 0.004) and the absence of STEC (p = 0.018) (Table 2). Lower CD4 cell counts were also associated with a longer duration of diarrhea (p = 0.0015) and older age (p = 0.011). Associations between organisms at CD4 count < 200/uL and < 50/uL were analyzed and none were found to be statistically significant.
The number of organisms detected in one sample was not associated with any specific demographic, including CD4 cell count, administration of HAART, age, duration of diarrhea or frequency of diarrhea. This was also the case when excluding B. hominis, D. fragilis and Aeromonas spp.
The risk of typhoid fever and non-typhoidal Salmonella invasive infections is highest in infants, young children, and young adults with underlying comorbidities, including severe anemia, malaria, malnutrition, and HIV infection. Moreover, recent reports from the international travelers agency showed that immunocompromised travelers, who usually follow the same itineraries of immunocompetent persons, visit countries at high risk of infections but the risk of developing travel-related diseases is five times higher if compared with that of immunocompetent persons.
However, data on groups at risk of acquiring typhoid infections are controversial and scant. Gordon showed that the immunological status cannot be associated with an increased risk or poor outcome. However, invasive diseases caused by non-typhoidal salmonellae are more frequently diagnosed in immunocompromised persons (e.g., persons with HIV/AIDS). Likewise, a study conducted in Africa did not find differences in HIV-positive patients and controls in the clinical presentation and outcomes of typhoid fever cases. In contrast, Gotuzzo and Colleagues found a rate of typhoid fever 25 times higher in HIV-positive patients than in the general population.
With the remarkable increased number of travelers from high-income countries during the last two decades, it was estimated that 1.9 million children traveled overseas every year from the United States; similarly, a significant increase of travelers (1.7 fold) was shown in Greece from 2004 to 2008. A high proportion of enteric fever cases was described in children aged 0–14 years (>26% in 2018), mainly attributed to tourism and VFR-travels. Zhou and Colleagues highlighted an increased rate of childhood enteric fever in a large tertiary care center in Canada during 1985–2013, with several cases caused by Salmonella paratyphi A and B and by bacterial strains resistant to first-line antibiotics. In Australia, 87% of the childhood cases were acquired mainly in Southeast Asia, with an annual increasing incidence from the period 2001–2005 (13 cases per year) to the period 2011–2015 (38 cases per year). Similar data were described in France, where children aged <18 years accounted for one-third of enteric fever patients, with 61% of the infections acquired in Africa.
Pre-travel counseling focused on hygiene and preventive measures could help reduce the risk of infection in individuals younger than two years, who cannot be immunized with the currently available vaccines.
The duck (Anas platyrhynchos) is one of the economically important poultry species as a source of meat, eggs and feathers. Ducks harbour most of the hemagglutinin (HA) and neuraminidase (NA) subtypes of avian influenza viruses that are currently known [2, 3] and serve as the principal natural reservoir host for influenza A viruses [2, 4–6]. Influenza A viruses maintained in wild aquatic birds have been associated with stable host switch events to novel hosts including mammals and domestic gallinaceous poultry leading to the emergence of novel influenza A viruses [7, 8].
To control the outbreaks of emerging or re-emerging viral diseases and prevent the transmission of viruses from the reservoir host, monitoring the virome status in the reservoir hosts is essential. Further, understanding the viral diversity in the poultry gut will improve the knowledge of enteric disease syndromes and the feed conversion efficiency of the poultry species. In recent years, next generation sequencing technology based viral metagenomics has provided a powerful tool for large-scale detection of known and unknown viruses existing in the reservoir host [11, 12]. Using this approach, known and novel viruses have been characterized from the enteric tract of turkey, bats, pigs, rodents, pigeon, ducks and ferrets. To obtain an unbiased measure of the viral diversity in the enteric tract of ducks, we deep sequenced viral nucleic acid isolated from cloacal swabs of 23 ducks collected from Bhoj wetland of Bhopal, the capital of the central Indian state of Madhya Pradesh. The present study revealed that the duck gut virome contained sequences related to a wide range of animal, insect, plant, and bacterial viruses. This study increases our understanding of the viral diversity present in the enteric tract of ducks. Further, this virome dataset provide a baseline faecal virome of the ducks and will be used as reference for identification of future changes in its virome composition, which may be associated with disease outbreaks or environmental changes.
The first ever report of RVs in poultry was from the USA (South Dakota) in 1977 from intestinal contents of turkey poults. In 1981, McNulty and coworkers isolated a virus (virus 132) from chickens of North Ireland. Despite having morphological and biochemical similarities to RVs, virus 132 was not antigenically related to any of the previously described RVs. This virus was categorized as atypical RV/non-group A RV. Based on comparative antigenic and nucleic acid analysis, Pedley et al. designated this atypical RV/non-group A RV as D/132, and proposed the extension of the number of RV groups by including RVD as new member. Later, Theil et al. (1986) found non-group A viruses from turkeys, which they called turkey rotavirus-like viruses (RVLVs). Likewise, RVLVs were also found in pheasants, and were later determined to be RVD. Since then, RVD has been reported frequently in turkeys, chickens, and pheasants, and sporadically reported in guinea fowls, partridges, quails, pigeons, ducks, etc.,. Up to now, the prevalence and clinical importance of RVD has been recognized for chickens and turkeys in various countries. European countries from where RVD infections have been reported include Scotland, Sweden, Germany, the UK, Italy, and the Netherlands. Apart from Europe, RVD has also been reported in chickens from the delta region of Egypt. In 2012, a report on RVD infection from Brazil noted an incidence rate of 53%. In the Asian continent, the first report on RVD was from Bangladesh. After two years, a second report came from India, with an incidence of 17.39%. A study conducted by Otto et al. (2012) revealed a combined prevalence of 65.9% from Europe and Bangladesh. Recently, in June 2017, nearly 32% of RVD shedding was observed in Nigerian birds. This study also suggested that host-permissive RVs may show cross-species transmission.
Although RVs cause enteric diseases in mammals and birds, they are often detected in otherwise healthy flocks, particularly when sensitive molecular diagnostic assays are used. Bezerra et al. (2014) reported the occurrence of RVD in apparently healthy asymptomatic chickens. Mixed enteric infections have also been reported. Saif et al. (1990) isolated RVD, AsTV (astrovirus), Salmonella, and small round viruses (18–24 nm in diameter) from an enteritis outbreak in turkey poults. RSS is a disease complex caused by many viruses, including reovirus, astrovirus, rotavirus etc. Four groups of AvRVs were recognized in flocks with RSS: avian RVA, RVD, RVF, and RVG. Otto et al. (2006) reported that RVD plays a major role in the pathogenesis of RSS in flocks with severe villous atrophy. A recently granted patent also confirms that among all of the AvRVs, RVD plays a major role in the pathogenesis of RSS. Earlier studies on rats have shown the capacity of AvRVs to disseminate and replicate in different organs, such as the liver, spleen, and pancreas; however, the mechanism by which RV escapes the gastrointestinal tract and reaches other organs remains unknown. Recently, the extra-intestinal presence of avian RVA in the pancreas and spleen of broilers with RSS has also been reported. However, there is no such report for RVD.
In India, the reports on RVD are very scarce. The first report on RVD came from central India in 2008, based on the specific electrophoretic migration pattern of RVD i.e., 5:2:2:2. In 2010, RVD was detected in western parts (Maharashtra) of India. The first sequence of confirmed RVD was reported in broiler chicks from northern India in 2013. Subsequently, in 2014, an RT-PCR was developed for detection of RVD, with a better detection limit (1.49 × 103 copies) than reported previously from Brazil. Using this reverse transcription-polymerase chain reaction (RT-PCR), a frequency distribution study was conducted in 2011–2012, which confirmed the existence of RVD in chickens from northern states of India with a 6.04% positivity rate. Using the same RT-PCR, recent (2012–2017) screening data revealed that this percent positivity in northern India has increased nearly fourfold over the last five years (Deol, unpublished data).
Only limited work has been carried out in India about coinfection of diarrheagenic bacterial and viral agents in porcine. Hence, the present study provides updated information and highlighted the significance of E. coli, Salmonella, Rotavirus, and Picobirnavirus as important diarrheagenic pathogens causing coinfection in piglets, where such data are scarce. In conclusion, it may be stated that coinfections with either of the four enteric bacterial and viral pathogens persist and associated with the piglet diarrhea in Northeast region of India, particularly with Picobirnavirus and S. Typhimurium. Higher prevalence is recorded from unorganized compared to organized farm and higher detection in crossbreed compared to the local indigenous pigs with higher detection during the summer season. This probably the first systematic study of the prevalence of the coinfection of important diarrheagenic bacterial and viral pathogens associated with piglet diarrhea in India.